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GGEXeiGHT DEPOSm 



Clinical Diagnosis 



A TEXT-BOOK 

of 

CLINICAL MICROSCOPY AND CLINICAL CHEMISTRY 

FOR MEDICAL STUDENTS, LABORATORY 

WORKERS, AND PRACTITIONERS 

OF MEDICINE 



BY 

CHARLES PHILLIPS EMERSON, A.B., M.D. 

LATE RESIDENT PHYSICIAN, THE JOHNS HOPKINS HOSPITAL; AND ASSOCIATE 

IN MEDICINE, THE JOHNS HOPKINS UNIVERSITY J PROFESSOR OF 

MEDICINE, INDIANA UNIVERSITY SCHOOL OF MEDICINE 



156 ILLUSTRATIONS 



FIFTH EDITION ENTIRELY REWRITTEN AND RESET 



PHILADELPHIA AND LONDON 
J. B. LIPPINCOTT COMPANY 






COPYRIGHT, I906, I908, 191 1, AND I913, BY J. B. LIPPINCOTT COMPANY 
COPYRIGHT, 1921, BY J. B. LIPPINCOTT COMPANY 



DEC i5iy2l 



Electrotyped and Printed by J. B. Lippincott Company 
The Washington Square Press, Philadelphia, U. S. A. 



CW.A653105 



Ox* ) 



To 
WILLIAM OSLER, M.D. 

IN GRATEFUL RECOGNITION 
OF THE MANY KINDNESSES 
RECEIVED BY A PUPIL AND 
ASSISTANT, THIS BOOK IS 
AFFECTIONATELY DEDICATED 
BY THE AUTHOR 



PREFACE TO THE FIFTH EDITION 

Ten years have passed since the last edition of this book appeared and 
seven years since its last copy disappeared from the market, periods of time 
which in personal history may seem short, since crowded by so many 
momentous events, but in the history of a text-book is only too long, since 
this Clinical Diagnosis now must present itself as entirely new and win 
whatever success it may, assisted little by the popularity it formerly 
enjoyed. The author desires to express his appreciation to the many 
friends who gave the previous editions such cordial reception and invites 
them now to examine critically this. 

As regards its contents this certainly is a new work. So much new 
material has appeared during the past ten years that a complete rewriting 
of the entire volume has been necessary and several new sections have been 
added, among them, those on serology, bacteriology, chemistry of the 
blood and of the spinal fluid. 

In presenting this edition to the inspection of the reader, the author 
takes the liberty to outline the general plan he has followed in its prepara- 
tion and to state the ideals which have influenced him. As in the case of 
the previous editions this book is not merely a manual for laboratory 
workers but is intended especially for medical students and for those 
practitioners of Internal Medicine whose conscience urges them to do their 
own laboratory work or at least personally to supervise and to interpret 
all the laboratory examinations made for each of their cases. The author 
feels that the present separation of laboratory and ward is an evil which 
cannot be too strongly condemned. No matter how successful a man may 
be he has no right to entrust his laboratory work entirely to others. Labora- 
tory assistants who can do the mechanical work and the cooperation of 
specialists in the medical science are necessary to the clinician but the 
conscientious man must regard the laboratory examinations as part of the 
general personal examination and must hold himself personally responsible 
for them all. The one who takes the history of the patient and makes the 
physical examination is the only one who can interpret correctly a labora- 
tory finding. Exactly identical reports may have quite different meanings 
in two different cases. He alone who knows the patient can interpret and 
evaluate a specimen under a microscope or in a test-tube and he often sees 
there that for the record of which no dotted line is provided on a laboratory 
blank but which will suggest further questions for the history and further 
physical examinations which may be of great value. The rather wide- 
spread and blind confidence which the past generation has placed in 
impersonal laboratory reports has brought Internal Medicine into a certain 
degree of disrepute. 



vi PREFACE TO THE FIFTH EDITION 

Medicine is an art, it cannot be a science. While it is not a science, it 
has found all the sciences very useful in its development as an art. The 
medical student should understand that the sum total of all laboratory 
activities is not and never can be internal medicine and that he as artist 
must try to see in each specimen something of the patient himself and 
interpret it in terms, not of chemistry, physics or biology, but of the 
suffering person. 

No attempt has been made to give a wide review of literature or to 
describe methods as yet untried. Those are recorded which the author 
and his associates personally have found valuable. This book is not the 
product of one man but of the teachers, associates and assistants of the 
Johns Hopkins and Indiana University Medical Schools with which it has 
been the privilege of the author to be associated. It is obviously impossible 
for him justly to acknowledge his indebtedness to the many who consciously 
or unconsciously have assisted him. He will, however, mention gratefully 
by name, among those who have assisted directly in the preparation of 
this edition, Dr. Virgil H. Moon, Prof. B. Bernard Turner, Dr. John R. 
Thrasher and Dr. Harry K. Langdon. 

Indianapolis, July i, 192 i 



PREFACE TO THE FOURTH EDITION 

The changes made in the preparation of this new edition are numerous 
and important. Our attempt has been to incorporate such important 
new contributions to the laboratory diagnosis of diseases as belong in a 
book of this, nature, and to omit methods which have been replaced by 
better ones. These omissions and the grouping of illustrations formerly 
in the text in plates have made it possible to preserve for the most part the 
paging of the third edition. The most extensive additions and alterations 
are those in the chapters on tuberculosis and lues. No effort has been 
spared to make this book accurate and practical. We would acknowledge 
with gratitude the helpful assistance of our many friends, among whom 
may be mentioned Dr. Clyde G. Guthrie of Baltimore, Dr. Roger Morris 
of St. Louis, Dr. John R. Thrasher of Indianapolis, Dr. Louis W. Ladd of 
Cleveland and Dr. H. S. Houghton of Wuhu, China. 

Charles P. Emerson. 
Indianapolis, 1913. 



PREFACE TO THE FIRST EDITION 

There have, during the past few years, appeared so many and such 
excellent text-books on clinical diagnosis, the clinical examination of the 
blood, the urine, or the gastric contents, that to add to this number one 
which covered the same ground in the same way as they, would seem a 
thankless undertaking, as well as an unpardonable misuse of energy. It 
is because the present work tries to cover this same ground in a different 
way, and one which will, we believe, commend itself to the medical, pro- 
fession, that we venture to offer it for inspection; we refer to the consider- 
ation of clinical laboratory work from the clinical rather than from the 
labor atory point of view. 

This book is based on the author's experience as physician in charge 
of the clinical laboratory, and instructor in medicine, of the Johns Hopkins 
Hospital and University. He has also had at his disposal all the clinical 
records of the ward cases for the seventeen years of this hospital's activity. 

Our course in clinical microscopy and chemistry extends over the 
eight months of the student's third year; two afternoons of three hours, 
and one of one hour, each week ; but much of the work is done out of class 
hours, as inspection of pages 447 and 485 will show. The subjects studied 
are the clinical examination of the blood, urine, sputum, stomach contents, 
feces, and various fluids, as ascitic, pleural, cerebrospinal, cyst contents, 
etc. In addition to this the student follows cases assigned him in the out- 
patient department. To those fitted for such work simple problems of 
research are given. The course is a laboratory one; specimens are provided 
each of the students. It is needless to say that with the eighty microscopes 
focussed on eighty specimens of a patient's blood, sputum, etc., the most 
of the interesting cells or other features will be found. The best were 
drawn by an artist always within call. The questions discussed in the 
following pages are for the most part those asked by the students during 
the class-work. The object of this course is not so much to impart knowl- 
edge as to raise the efficiency of the student. It is not a course in chemistry 
and microscopy, but in these applied to the study of a patient ; not in physi- 
ology, but in pathology. With the methods of chemical and biological 
work, with the normal findings, they are already familiar. Chemistry, 
inorganic and organic, qualitative and quantitative, is required for admis- 



x PREFACE TO THE FIRST EDITION 

sion to the school; the normal blood they have studied in the anatomical 
laboratory ; normal urine and gastric contents, in the laboratory of pr^sio- 
logical chemistry. We take this knowledge for granted as a foundation 
for the study of pathological bloods, urines, etc., paying particular atten- 
tion to the clinical significance of these findings. At the same time the 
students are required to practise the best methods in e very-day use, not 
only until they understand them, but until they can accurately use them. 
It is the practical "use of a determination or examination which is empha- 
sized. If approximate methods will do, they are used; if accurate methods 
are necessary, accurate work must be done, whatever the cost in time. 
To use an approximate method well is far better than to employ a more 
exact, laborious one poorly; to do approximate work is not always easy 
and requires practice; to be able to do accurate work well is also required 
of our students. Practice, experience, an exact knowledge, first of the 
possibilities in a method, second, and just as important, his own accuracy 
in the use of that method — these it is the duty of the clinical laboratory 
to give a student. Above all, he should train his common sense so that, 
using his eyes, nose, ears, and tongue, he can get results for which another 
man would apply elaborate methods. 

The author has been careful not to include new untried methods, for 
of these but a small number will last, and a text-book should contain noth- 
ing as yet not well tested by friends and foes. It is the introduction of 
" new methods " which renders some books even dangerous to the man 
who buys but one. 

We do not claim that with this book alone the student can study 
clinical microscopy. No subject in medicine is broader or requires more 
reference books, for some of the hardest chemical problems will at times 
confront him, and to interpret the various artefacts and accidental findings 
of the microscope would require a vast experience in microscopy, and a 
knowledge of zoology, botany, and mineralogy as broad as is the realm 
of science. For who knows what infusoria, what diatom, desmid, or other 
protophyte, the ovum of what parasite, the wing of what insect, the leg 
of what fly, the tissue of what plant, the fiber of what meat, the seeds of 
what berry or fruit, may be found in sputum, stomach contents, urine or 
feces, from the food, tap-water, or the contaminations from dirty vessels, 
or from the dust of the air ? To be wise in the points of differential chemical 
and microscopical diagnosis is splendid; but to recognize artefacts and 
extraneous matter, the stumbling-blocks in diagnosis, that is the true test 



PREFACE TO THE FIRST EDITION xi 

of the clinical laboratory worker, and this ability is gained by wide experi- 
ence alone. 

The function of the clinical laboratory worker is to aid the ward worker. 
The findings of the former are seldom conclusive, and must be interpreted 
in the light of the ward findings ; especially is this true now that functional 
diagnosis is the goal. The writer can only give to the reader who has 
aspirations to be a clinical chemist and microscopist the advice in substance 
which one of Germany's greatest clinical chemists gave him when the latter 
regretfully left the little Swiss laboratory which had been such a pleasant 
home: the clinical chemist must be first a good clinician and second a 
chemist ; he should remember that even from the laboratory point of view 
his stethoscope is of more importance than his microscope, his percussion 
finger than his whole outfit of chemical apparatus. 

In conclusion, we wish to express our indebtedness to Doctor Osier for his 
encouragement and aid during the progress of this work, and for his hearty 
cooperation in placing at our disposal the records of the medical wards; 
and to the assistants and students of this clinic, for whose aid I am very 
grateful, and who are too many to mention by name except Dr. Thomas 
R. Boggs, whose suggestions and criticisms have been so valuable. 

I take this opportunity to thank the artists who have done much 
beautiful work for me — Messrs. F. S. Lockwood, Hermann Becker, Max 
Brodel, and Mrs. Ruth Huntington Brodel, whose excellent half-tone and 
pen-and-ink drawings must be recognized by the lack of her signature. 

Charles P. Emerson. 
Johns Hopkins Hospital, 1906. 



INTRODUCTION 



The clinical laboratory has two special functions in the medical 
school, — in it the student learns the application of physical and chemi- 
cal methods in the study of disease, and in it researches are conducted 
on the innumerable problems concerning etiology, diagnosis, and treat- 
ment. Forming an essential part of the hospital-half of a school, it 
should be close to the wards and so arranged as to have ample facilities 
for the students and for the house physicians and others doing special 
work. It should be in charge of a man resident in the hospital, 
familiar with the routine of the clinic, and in close daily touch with 
his chief and with the assistants. The expenses should be shared 
equally by the hospital and the medical school. Into the details of 
organization I will not enter, but the director of such a laboratory 
should, if possible, have assistants thoroughly trained in bacteriology, 
physiological methods, and physiological chemistry. 

In 1896, through the kindness of two ladies, a special clinical 
laboratory was built for the students of the Johns Hopkins Medical 
School, which was enlarged tw T o years ago when the new clinical 
building was erected. On each of the two floors about fifty students 
are accommodated and there are rooms adjacent for special workers 
and for the assistants. Dr. Jesse Lazear was at first in charge, and 
under Dr. Thayer's direction the well-known researches of Macallum 
and Opie and of Lazear himself on malaria were carried on. In 1900, 
after Dr. Lazear went to Cuba, we were fortunate enough to have Dr. 
Charles P. Emerson take charge of the laboratory, and to him the 
medical school is deeply indebted for the organization of clinical 
laboratory courses of the most thorough and scientific character. 

In medical education the all-important problem is to give a man 
the knowledge he can use. In our modern system much of the train- 
ing is rendered ineffective, as it has not been sufficiently prolonged 
to become part of a man's intellectual or bodily mechanism. A brief 
course of six weeks on any practical subject is almost useless and in 
some may be positively dangerous. When possible, an orderly 
sequence should be followed, so that the work of each year 
shall supplement that of the preceding. In the seven-year course 
laid down by the Johns Hopkins University a thorough laboratory 
training in biology, physics, and chemistry is given before the profes- 



xiv INTKODTJCTION 

sional work begins, so that a man enters the medical school proper 
with a practical knowledge of scientific methods and of the use of 
instruments of precision. In his first year of the medical curriculum 
the courses in histology and physiology and in the second year those 
in physiology, bacteriology, physiological chemistry, and pathological 
histology give him an insight into the structure and functions of the 
body, and he becomes thoroughly familiar with the use of all instru- 
ments of precision. In the third and fourth years in the hospital side 
of his education, for which the previous ones have been a preparation, 
he must have opportunities to carry on his practical work, and these 
the clinical laboratory affords. A student who has been interested 
in the mysteries and mechanism of cardiac rhythm in the physiological 
course should be able to take the pulse and heart tracings of the first 
case of mitral disease that he meets in the out-patient department, and 
the means should be afforded him to pass without a jar from the 
normal to the abnormal, — without, indeed, appreciating that there is 
any difference in the method of approaching the problems involved. 
So too a student should be able at once to attack his first case of 
diabetes as a problem in carbohydrate metabolism, fully prepared by 
previous study to approach it on the clinical side. 

If the curriculum were not so full, a student could gradually work 
out for himself, as the patients came under observation, every detail 
in the application of scientific methods to clinical study, but it is found 
more convenient to group them together and present in orderly 
sequence the subjects for study. Concurrently with the systematic 
instruction in the out-patient department which forms a large part 
of the work of the third year, a course on microscopical and chemical 
methods is given, and each man has his own place in the laboratory 
at which he may work throughout the year. This book is the outcome 
of the work by Dr. Emerson and his students in this course during 
the past five years. Not only does it represent the results of a very 
large number of careful observations made in the laboratory, but 
an analysis of many important groups of cases in the wards, so that 
it illustrates the experience of the medical clinic of this hospital so far 
as it relates to microscopical and chemical methods of diagnosis. 
The work will be found a comprehensive and trustworthy guide in 
all the details of laboratory work. 

But the aim of a training such as this book implies is to send 
out into practice men able to give patients the benefit of modern scien- 
tific methods in the diagnosis and treatment of disease — men who use 



INTBODUCTION xv 

the microscope, who examine sputum, and who use the stethoscope, 
and who can do the routine urine and blood work with confidence. 
The men to study a book of this kind are the young practitioners who 
are keeping up the practical knowledge obtained in the medical school, 
and who appreciate a small laboratory as the most valuable stock-in- 
trade. As a practitioner becomes more and more engaged, he can 
hand over to an assistant the laboratory side of the work, but it is 
surprising how much can be done even by the busiest of men if the 
will is there and if the methods have once been thoroughly mastered. 

William Osler. 

January 30, 1906. 



CONTENTS 



PAGE 

Introduction xiii 

CHAPTER I. 

THE SPUTUM. 

Introduction I 

General Features I 

Amount of Sputum 2 

Consistency of Sputum 3 

Reaction of Sputum 3 

Character of Sputum 3 

Color of Sputum 3 

Air of Sputum 5 

Layer Formation of Sputum 5 

Odor of Sputum 5 

Macroscopic Constituents of Sputum 6 

Microscopic Constituents of Sputum 11 

Plant Parasites in Sputum 18 

Animal Parasites in Sputum 50 

Chemical Examination of Sputum 53 

Sputum in Disease : Pulmonary Tuberculosis 54 

Pneumonia , 60 

Bronchopneumonia 65 

Influenza 65 

Whooping-cough 66 

Glanders 66 

Asthma 66 

Acute Bronchitis 68 

Chronic Bronchitis 70 

Bronchiectasis 74 

Gangrene of the Lung 77 

Aspirated Foreign Bodies 78 

Abscess of the Lung 78 

Perforating Empyema . 80 

Perforating Serous Pleurisy 80 

Edema of the Lungs 80 

The Albuminous Sputum of Thoracentesis , 81 

Hemoptysis 81 

Sputum in Hemorrhagic Infarction of the Lungs 83 

Chronic Passive Congestion of the Lung 83 

Malignant Disease of the Lungs 83 

Syphilis of the Lung 84 

Pneumokoniosis 85 

Sputum in Diphtheria 85 

Vincent's angina 87 

xvii 



xviii CONTENTS 

CHAPTER II. 

THE URINE. 

PAGE 

General Characteristics 88 

The Collection and Preservation of Urine 88 

The Amount of Urine * 89 

The Specific Gravity of Urine ~ 92 

The Color of Urine 94 

The Pigments of Urine 95 

The Odor of Urine 99 

The General Appearance of Urine 100 

The Reaction of Urine „ 100 

The Nitrogenous Bodies of the Urine 104 

Nitrogen 104 

Urea 109 

Uric Acid 114 

Purin Bases 120 

Ammonia 122 

Creatinin 125 

Creatin 128 

Oxyproteinic and Alloxyproteinic Acids 128 

The Inorganic Acids and Bases of Urine 129 

Chlorides 129 

Phosphates 133 

Sulphates 136 

Thiosulphuric Acid '. 140 

Hydrogen Sulphide 140 

Sulphocyanic Acid '. 140 

Carbonates 140 

Calcium and Magnesium 140 

Sodium and Potassium 142 

Iron 142 

Lead 143 

Arsenic 143 

The Pigments of the Urine 143 

Indoxyl Sulphate 143 

Phenol 146 

Skatoxyl-Sulphate 147 

Indigo-red 147 

Paracresol and Phenolsulphuric Acid 148 

Pyrocatechin 148 

Hydrochinon 148 

Potassium Iodide ' 148 

Bile Pigments 148 

Bilirubin 149 

Biliverdin , . . 150 

Hydrobilirubin 150 

Bilifuscin 150 

Biliprasin 150 

Cholecyanin 150 

Choletelin 150 

The Reducible Body of Stokvis 151 

Test for Bile Pigments 151 



CONTENTS xix 

The Pigments of the Urine page 

Melanin- Melanogen 155 

Rosenbach's Reaction . 155 

Bile Acids 155 

Diazo Test 157 

Ferments in the Urine 159 

Pepsin 159 

Diastatic Ferment 160 

Lipase 160 

Amylase of the Urine 160 

Carbohydrates and Allied Bodies in the Urine 162 

Glycosuria 164 

Levulose 181 

Lactose 182 

Pentoses 183 

Inosite 186 

Glycogen (Erythrodextrin) 186 

Animal Gum (Landwehr) 187 

Laiose 187 

Maltose 187 

Isomaltose 187 

Melituria 187 

Acetone 187 

Diacetic Acid 192 

/3-Oxybutyric Acid 194 

Diabetes Mellitus 198 

Diabetes Insipidus 205 

Glycuronic Acid 206 

Alkaptonuria 207 

Homogentisinic Acid 208 

Uroleucinic Acid 208 

Fat in the Urine 209 

Proteids in the Urine 210 

Albumin Tests 210 

Proteids Present 217 

Serum Albumin 218 

Serum Globulin 218 

Euglobulin, Nucleo-Albumin, Mucin, Morner's Body 219 

Nucleohiston 223 

Fibrinuria: Fibrinogen, Fibrinoglobulin 224 

Albuminuria without Definite Renal Lesion 224 

Physiological Albuminuria 224 

Functional Albuminuria 225 

Albuminuria of the New-born 227 

Albuminuria of Women in Labor 227 

Albuminuria of Adolescence 227 

Cyclic Albuminuria 228 

Hypostatic Albuminuria 230 

Albuminuria Minima 230 

Intermittent Albuminuria 230 

Traumatic Albuminuria 23 1 

Febrile Albuminuria 23 1 

Hematogenous Albuminuria 23 1 



xx CONTENTS 



Proteids Present page 

Nervous Form of Albuminuria 23 1 

Albuminuria with Definite Renal Lesions 232 

Nephritis 232 

Hetero-Albumosuria, Bence- Jones Body 233 

Albumosuria, Deutero-Albumosuria, Peptonuria 235 

Hematuria _ 237 

Hemoglobinuria 238 

Methemoglobin 242 

Hematoporphyrin 242 

Sediments 243 

Preservation of the Urine 243 

Unorganized Sediments 245 

Urates and Uric Acid 245 

Phosphates and Carbonates 248 

Oxaluria 250 

Calcium Sulphate 252 

Hippuric Acid 253 

Hetero-Albumose 253 

Xanthin 253 

Hematoidin (Bilirubin) 253 

Indigo 253 

Melanin 253 

Hemoglobin 253 

Cholesterol 253 

Leucin and Tyrosin 254 

Cystin 256 

Diamines 257 

Scheme of Sediments 258 

Chyluria 259 

Lipuria 260 

Organized Sediments 261 

Mucous Sediment 26 1 

Epithelial Cells 261 

Hemosiderin in Urine 261 

Casts 264 

Epithelial 264 

Granular 265 

Fatty 266 

Waxy. 266 

Hyaline, Colloid, Glassy 267 

Blood 267 

Hemoglobin 267 

Pus 267 

Cylindroids 267 

Combined Casts; Bacterial Casts, Urate Casts 268 

Pseudo-Casts 269 

Diagnostic Importance 270 

Testicular Casts 273 

Gonorrheal Threads, Clap Threads 273 

Tissue Fragments 274 



CONTENTS xxi 

PAGE 

Organized Sediments 

Pus-Cells 274 

Red Blood-Corpuscles 276 

Concretions 276 

Renal and Bladder Stones: Urate and Uric Acid 276 

Table of 277 

Oxalate 278 

Phosphate 278 

Carbonate 278 

Cystin 278 

Xanthin 278 

Fatty 279 

Indigo 279 

Albumin . 279 

Bacteriology of the Urine 279 

Technic of Obtaining Specimens 279 

Bacterioscopic Examination 280 

Bacterial Stains 280 

Spore Staining; Flagella Staining 282 

Cultural Method 284 

Media 285 

Organisms that may be found in the Urine 286 

Bacillus Coli Communis 286 

Bacillus Typhosus 287 

The Paratyphosus Group 287 

Bacillus Lactis Aerogenes 288 

Bacillus Alkaligenes 288 

The Proteus Group 288 

Bacillus Pyocyaneus 289 

Bacillus Aerogenes Capsulatus 289 

Bacillus Tetani 290 

Septicemia 290 

Infectious Nephritis 29 1 

Acute Pyelitis 29 1 

Cystitis 291 

Bacteriuria 294 

Infection of Urethra and External Genitals 294 

The Gonococcus 294 

Acute Anterior Urethritis 295 

Posterior Urethritis 296 

Non-specific Urethritis 297 

Bacteriorrhea 297 

Prostatitis 297 

Micro-organisms of External Genitalia 300 

Bacillus Ulceris Cancrosi 300 

Treponema Pallidum 300 

Spirocheta Refringens 302 

Treponema Microdentium 303 

Treponema Macrodentium 303 

Treponema Mucosum 303 

Treponema Calligyrum 304 

Yeasts, Moulds, Sarcinas 305 



xxii CONTENTS 



PAGE 



Animal Parasites 306 

Prostatic Fluid 308 

Functional Renal Diagnosis \ . 309 

Physiochemical Tests 310 

Methods Using Normal Products of Metabolism 311 

Test Meals 312 

Renal Permeability 318 

Phenolsulphonephthalein Test 319 

Phlorizin Test 323 

Value of Tests . 324 

Disturbed Renal Function in conditions without Renal Disease . , 324 

Diseases of the Kidneys , 324 

Albuminuria 325 

Acute Nephritis 327 

Nephritis Hemoglobinuria „ 328 

Subacute Nephritis 329 

Chronic Nephritis 332 

Uremia 333 

Unilateral and Nephritis 335 

Renal Atrophy 335 

Congenital Cystic Kidney 335 

Suppurative Nephritis 336 

Cancer of the Kidney 336 

Tuberculosis of the Kidney 336 

Infarction of the Kidney 337 

Pyelitis, Pyelonephritis 337 

Hydronephrosis, Pyonephrosis, Uronephrosis 338 

Renal Calculus 338 

Parasitic Diseases 338 

CHAPTER III. 

THE STOMACH CONTENTS. 

The Vomitus 339 

The Fasting Stomach 341 

Test Meals 342 

Gastric Acidity 344 

Total Acidity 345 

Free Hydrochloric Acid 347 

Hydrochloric Acid Deficit 348 

Total Hydrochloric Acid 348 

Physiology of the Gastric Secretion 349 

Diagnostic Value 349 

Fractional Determination of Gastric Secretion 351 

Sahli's Desmoid Reaction 354 

Pepsin 354 

Fat-vSplitting Ferment 356 

Rennin 356 

Products of Gastric Protein Digestion 356 

Starch Digestion 357 

Lactic Acid 357 

Other Organic Acids 359 



CONTENTS xxiii 

PAGE 

The Vomitus: Bases of Gastric Juice 359 

Fermentation 359 

Gastric Sediment 360 

Microscopic Examination 363 

Motility of the Stomach 364 

Hyperacidity 367 

Hypersecretion 367 

Nervous Dyspepsia 369 

Acute Gastritis 369 

Chronic Gastritis 370 

Mucus 371 

Atrophy of Mucosa 372 

Uicer of Stomach 373 

Syphilis of the Stomach 374 

Cancer of Stomach 375 

CHAPTER IV. 

THE INTESTINAL CONTENTS AND THE FECES. 

Pancreatic Fluid 382 

Trypsin 382 

Fat-Splitting Ferment 382 

Diastase 382 

Bile from Duodeum 383 

Motility of Intestine 385 

Test Meals 385 

Examination of Stools 386 

The Constituents of Normal Stools 387 

The Reaction of the Stools 387 

The Frequency of the Stools 387 

The Consistency and Form of the Stools 387 

The Color of the Stools 389 

Acholic Stools 389 

Fatty Stools 390 

Mucus in the Stools 392 

Blood in the Stools 393 

Pus in the Stools 395 

Undigested Food in the Stools 396 

Microscopy of the Stools 397 

Macroscopical Examination of the Stools 398 

Concretions 398 

Tumor Fragments 401 

Animal Parasites 401 

Plant Parasites 418 

Stools in Disease 420 

Typhoid Fever 420 

Asiatic Cholera 1 422 

Dysentery, Rectal Diarrhea, Cancer of the Rectum 423 

Amebic Dysentery 423 

Pancreatic Disease 424 

Permanent Mounts of Small Worms 426 



xxiv CONTENTS 



CHAPTER V. 

THE BLOOD 



PAGE 



Obtaining the Blood 428 

Specific Gravity 429 

Dried Residue _ 43 1 

Coagulation 43 1 

Viscosity 437 

Hemoglobin 439 

Fresh Blood 445 

Red Cells '. . . 445 

Degenerations 448 

Leucocytes 450 

Hemokonien Granules , 451 

Platelets 452 

Fibrin 452 

Blood Counting, red cells 452 

Leucocytes 461 

Blood Smears 462 

Fixing Methods 464 

Stains 466 

Red Cells 473 

Shape 473 

Structure 473 

Size 474 

Staining Properties 475 

Granules of Red Cells 477 

Number of Red Cells 482 

Physiological Variations 483 

Drugs and Therapeutic Measures 486 

Pathological Conditions 487 

Polycythemia 487 

Resistance of Red Cells 489 

Color Index 490 

White Cells 491 

Granules 491 

Classification 494 

Differential Counting 498 

Bone-Marrow 500 

Nucleated Red 500 

Origin of Red-cells 503 

Origin of Leucocytes 504 

Fetal Blood 5°7 

Leucocytosis • • 5°8 

Digestion 5°9 

New born 5 1 1 

Inflammatory 5 * 2 

Pseudoleucocytosis 5 : 7 



CONTENTS xxv 



PAGE 



Leucocytosis 

Malignant Disease 517 

Post-hemorrhagic 518 

Agonal 518 

Medicinal 518 

Mixed 519 

Mastzell 519 

Endothelial 519 

Lymphocytosis 519 

Leucopenia 521 

Eosinophilia 52 1 

Iodophilia 524 

Blood Platelets 525 

Chemistry of Blood 528 

Hemoglobinemia 528 

Methemoglobinemia 528 

Sulph-hemoglobinemia 529 

Bilirubin and Urobilin in Blood 529 

Reaction of Blood 53 1 

Chlorides 538 

Nitrogenous Bodies 539 

Nitrogen 539 

Urea 541 

Ammonia 542 

Uric Acid 543 

Creatinin 547 

Creatin 548 

Glucose in Blood 549 

Diastatic Activity 554 

Lipoids 555 

Cholesterol 558 

Bacteriology of Blood 561 

Streptococcus group 563 

Typhoid Bacilli 563 

Paratyphoid Bacilli 563 

Agglutination Phenomena 564 

Gruber-WidalTest 564 

Other Agglutinations 567 

Opsonins 567 

Complement Fixation 569 

The Wassermann Test 575 

Gonococcus Fixation 584 

Tuberculosis . 586 

Isohemagglutinins 588 

Anemia 590 

Secondary 592 

Simple Primary 602 

Pernicious 602 

Chlorosis 612 

Aplastic 615 



xxvi CONTENTS 

PAGE 

Leukemia 6 1 6 

Splenomyelogenous 617 

Lymphatic 623 

Acute 625 

Mixed 628 

Pseudoleukemia 628 

Hodgkins' Disease ± 628 

Tuberculosis of Lymph-Glands 628 

Leukanemia . . 629 

Blood in Acute Diseases 629 

Tuberculosis 634 

Typhoid Fever 638 

Pneumonia, etc 640 

Diseases of Children 645 

Chronic Diseases 647 

Diabetes Mellitus 647 

Malignant Disease 648 

Renal Disease 654 

Diseases of Liver, etc 656 

Value of Blood Examination 658 

Malaria 659 

Fresh Blood 660 

Tertian 660 

Quartan 662 

^Estivo-Autumnal 664 

Cycle in Mosquito 665 

Stained Specimens 667 

Tertian 667 

Quartan 669 

^Estivo-Autumual 669 

Trypanosomiasis 67 1 

Piroplasmosis : 672 

Filariasis 673 

Relapsing Fever 677 

CHAPTER VI. 

CEREBROSPINAL FLUID. 

Cerebrospinal Fluid 678 

Glucose 679 

Urea 680 

Proteins 681 

Tests for Globulins 681 

Saline Constituents . . . . f 683 

Cholin 683 

Cytology 684 

Bacteriology 685 

Colloidal Gold Test 685 

In Diseases 688 

Uremia 688 

Diabetes Mellitus 688 

Chorea 688 



CONTENTS xxvii 

PAGE 

Cerebrospinal Fluid in Diseases 

Epilepsy 688 

Mental Diseases 688 

Hydrocephalus 688 

Cerebral Hemorrhage 688 

Brain Tumors 688 

Compression of Cord 689 

Encephalitis 689 

Meningism 689 

Meningitis 689 

Epidemic 689 

Pyogenic 690 

Tuberculous 690 

Lues 691 

Poliomyelitis 692 

CHAPTER VII. 

EXAMINATION OF VARIOUS FLUIDS. 

Various Fluids : Specific Gravity 693 

Proteids 693 

Fat, etc 694 

Milk 699 

Transudates and Exudates 699 

Peritoneal Fluid 699 

Pleural Fluid 700 

Empyema 703 

Pericardial Fluid 704 

Synovial Fluid 704 

Chylous and Chyliform 705 

Ovarian Cysts 706 

Hydrocele 708 

Spermatocele 708 

Tophi of Gout 709 

Urea Frost 709 



LIST OF ILLUSTRATIONS 



Plate I. Blood-cells. Ehrlich's stain 494 

" II. Leucocytes, platelets, and Trypanosoma, Hastings' stain 524 

" III. Parasites of tertian fever and quartan fever 660 

" IV. Parasite of aestivo-autumnal fever 664 

" V. Malaria (stained) 666 



FIG. PAGE 

1. Curschmann's spiral 7 

2. Free central fiber of a Cursch- 

mann's spiral 7 

3. Spiral thread of mucus from spu- 

tum 8 

4. Extraneous matter common in 

the sputum 12 

5. Epithelial cells found in sputum 12 

6. Elastic tissue from tuberculous 

sputum 14 

7. Elastic tissue from tuberculous 

sputum showing alveolar ar- 
rangement 14 

8. Fatty acid crystals in sputum ... 15 

9. Lep to thrix form in sputum 16 

10. Elastic tissue in sputum from 
food 17 

11. Micrococcus aureus 19 

12. Streptococcus pyogenes 20 

13. Bacillus tuberculosis 23 

14. Bacillus influenzae 35 

15. Bacillus diphtherias 38 

16. Smear from case of Vincent's 

angina 41 

17. Balantidium Coli photographed 

in Stool] 44 

18. Mucor mucedo 46 

19. Aspergillus fumigatus 47 

20. Aspergillus flavus 48 

21. Penicillium glaucum 50 

22. Sediment from echinococcus 
cyst.. 51 

23. Fragment from echinococcus 
cyst wall 52 

24. Egg of Paragonimus wester- 

mani 53 

25. Fibrin cast from a case of double 

pneumonia 63 

26. Kjeldahl apparatus for nitrogen 

determination 107 

27. Folin's apparatus for urea, am- 

monia, etc. determination. ... 113 

28. Schlosing's apparatus for am- 

monia determination 123 

29. Folin's apparatus for ammonia 

and acetone determination .... 124 

30. Apparatus for determining the 

melting point of crystals 173 

31. Half -shadow saccharometer 178 

32. Fields of a saccharometer 180 

33. Iodoform crystals 189 

34. The horismascope • 212 



I FIG. 

35- 
36. 
37. 
38. 
39- 
40. 

41. 

42. 

43- 
44. 

45- 

46. 

47- 
48. 

49. 
50. 
5i. 

52. 

53- 

54- 

55- 
56. 
57- 
58. 
59- 
60. 

61. 
62. 

63. 
64. 

65. 
66. 

67. 
68. 
69. 
70. 

71. 
72. 

73- 
74- 
75. 
76. 



Esbach's albuminometer 216 

Hemin crystals 241 

Ammonium biurate crystals .... 246 

Uric acid crystals 247 

Triple phosphate crystals 248 

Atypical triple phosphate cry- 
stals 248 

Dicalcium phosphate crystals . . . 249 
Sheaves of jcalcium phosphate 

needles 249 

Calcium phosphate crystals 249 

Calcium carbonate dumb-bells . . 250 
Calcium oxalate crystals and 

spheres 251 

Calcium oxalate crystals 251 

Various crystals 252 

Hematoidin, leucin, tyrosin, 

xanthin 253 

Cystin crystals 257 

Epithelial cells from urethra .... 262 

Epithelial cells from urine 263 

Epithelial cast and cells, pseudo 

pus-cast, etc 264 

Coarsely and finely granular 

casts 265 

Waxy casts 265 

Epithelial, fatty, and pus casts . . 266 

Hyaline casts 267 

Blood-cast 267 

Cylindroids 268 

Pseudo-casts 269 

Spread of pus containing gono- 

cocci 295 

Prostatic fluid 298 

Prostatic fluid 298 

Mucus mass full of spermatozoa . 299 

Cells in prostatic fluid 299 

Treponema pallida and Spiro- 

cheta refringens 303 

Protophytes, etc. often found 

in tap water 304 

Blastomycetes in urine 305 

Eggs of Eustrongylus gigas 306 

Schistosoma hematobium 307 

Strauss' funnel for lactic acid 

tests 358 

Sarcina ventriculi and yeast cells 363 

Fats and soaps in stools 390 

Fatty acid crystals 39 1 

Charcot-Leyden crystals 398 

Pseudo-eggs in stools 398 

Cells in stools 398 



XXX 



LIST OF ILLUSTRATIONS 



FIG. PAGE 

yj. Spines forming the "down" of 

fruits 399 

78. Ameba coli 401 

79. Trichomonas vaginalis 404 

80. Lamblia intestinalis 405 

81. Balantidium coli 406 

82. Ameba coli 407 

83. Trichina spiralis 408 

84. Eggs of Trichocephalus dispar 

and of Ascaris lumbricoides . . . 408 

85. Eggs of Tyroglyphus siro 408 

86. Oxyuris vermicularis 409 

87. Ankylostomum duodenale 410 

88. Caudal bursa of Uncinaria Amer- 

icana 411 

89. Eggs of Uncinaria duodenalis ... 411 

90. Head of Uncinaria americana . . . 412 

91. Head of Uncinaria duodenalis. . . 412 

92. Caudal bursa of Uncinaria duo- 

denalis 412 

93. Larva of Uncinaria americana. . . 413 

94. Dicrocelium lanceolatum 415 

95. Links of Tenia saginata 415 

96. Eggs of Tenia saginata 416 

97. Links of Tenia solium 416 

98. Tenia solium — head, link and 

egg 417 

99. Head of Tenia saginata 417 

100. Hymenolepis nana 418 

101. Bothriocephalus latus 419 

102. Egg of Schistosoma haemato- 

bium 419 

103. Egg of Schistosoma haemato- 

bium 420 

104. Bacillus bifidus 420 

105. Method of making cover-glass 

preparations 428 

106. Bogg's modification of Russell- 

Brodie coagulometer 435 

107. Diagrams of the movement of 

cells in the coagulometer 435 

108. Hiss' viscosimeter 437 

109. Mixing pipette for the Meischer 

hemoglobinometer 439 

no. Color-prism for the Meischer 

hemoglobinometer 440 

111. Pipette for the Fleischl hemo- 

globinometer 441 

112. Meischer's modification of the 

Fleischl hemoglobinometer.... 441 

113. Gowers' hemoglobinometer 442 

114. Sahli's hemometer 442 

115. Dare's hemoglobinometer 443 

116. Pipette of Dare's hemoglobi- 

nometer 443 



FIG. 

117. 
118. 
119. 

120. 
121. 
122. 
123. 
I24. 

125. 
126. 



127. 

128. 



I29. 
130. 

131. 
132. 
133. 
134. 



135- 
136. 

137. 

138. 
139. 
140. 
141. 
142. 

143. 
144. 

145- 
146. 

147. 

148. 
149. 

150. 

151- 



PAGE 

Fresh blood-cells 446 

Thoma-Zeiss blood-counter. .... 453 

Wynn's roller device 454 

Buerker's counting chamber 455 

Scheme of ruled slide ,456 

Arm of hematocrit 460 

Field of ruled slide. 462 

Nucleated red cells of fetal 

blood 462 

Blood platelets 462 

Apparatus for saturating blood 

plasma with C0 2 463 

C0 2 apparatus 464 

Apparatus for N determin- 
ation 465 

Blood culture apparatus 466 

Tubes for collection of blood 

for serum diagnosis 467 

Tube for diluting serum 471 

Widal test, negative result 471 

Widal test, positive result 472 

Hasmameeba leukaemiae magna 
et parva; large granular cell 

of bone-marrow 474 

Intestine of an infected mos- 
quito with oocysts 475 

The development of the ma- 
laria parasite in the mosquito's 

stomach 476 

Culex and Anopheles mos- 
quitoes 477 

Heads of mosquitoes 477 

Filaria bancrofti. 478 

Microfilaria with sheath 478 

Larva in mosquito's proboscis. . . 479 

Spirochete obermeieri 480 

Colloidal Geld curves 481 

Smear of spinal fluid of a case of 
epidemic cerebrospinal men- 
ingitis 482 

Smear of spinal fluid of a case of 
meningitis due to Diplococcus 

lanceolatus 482 

Smear of spinal fluid of a case of 
meningitis due to Bacillus 

influenzae 485 

Cells from a pleural fluid 486 

Fatty acid crystals from an ovar- 
ian cyst 489 

Cholesterol crystals 490 

Sodium biurate crystals 491 

Chart I Duodenal ulcer 
Chart II Duodenal ulcer 



CLINICAL DIAGNOSIS 



CHAPTER I 

THE SPUTUM 

Introduction. — The examination of the sputum is fast becoming a lost 
art. The discovery of a few specific microorganisms and the hope of finding 
many more, the American desire for a "sure" test and his enthusiastic 
study of new subjects, the "latest things" in medicine, all have led him 
gradually to abandon a great and valuable fund of knowledge. Today a 
sputum examination seldom means more than the search for Bacillus tuber- 
culosis and that by a State laboratory worker. That the average clinician 
now does not even glance at fresh sputum is proven by the presence on the 
market of red paper sputum cups only; red, so that the patient may not 
notice that he is expectorating blood ; red of such tint that the doctor can- 
not see the many delicate shades and characteristics of sputum which 
would have taught our fathers in medicine much of interest and value 
concerning the patient. There certainly are many other diseases of the 
lungs than tuberculosis and even in that one disease our duty involves 
not only an accurate diagnosis but an accurate prognosis as well and that 
we may be able to do this we must follow our case in its changes from day 
to day. For this reason a wise interpretation of the daily sputum changes 
is more important than a report from a State laboratory or a rontgeno- 
logical department. Our fathers, who never saw a germ, could diagnose 
and follow some pulmonary cases better than can we. They would not 
have made some of the mistakes which now humiliate us. The variety of 
colors, of physical and chemical characteristics and of structures, which 
the sputum may present or contain, is bewildering. Some of these are very 
important, more are negligible ; which are which, the clinician should 
know. He should not mistake bacteria in chains for elastic tissue, nor 
myelin globules for blood-cells. For a would-be Bizzozero to have expen- 
sive pictures of ordinary starch granules drawn, confident that a newly- 
discovered parasite will bear his name, probably means that when a medical 
student he did not study fresh sputum with enthusiasm. 

GENERAL FEATURES 

The sputum is, strictly speaking, that substance or mass of substances 
which is expectorated. The term covers not only the secretions from the 

1 



2 CLINICAL DIAGNOSIS 

respiratory passages but also any constituents added to these from the 
esophagus, nose and mouth, or, through perforation into these, from any 
neighboring organ. 

The normal person should theoretically have no sputum and yet persons 
who live in an atmosphere laden with dust may every morning expectorate 
lumps, often as large as a cherry, which are tough, elastic, so translucent 
that they resemble boiled sago (due to myelin) and gray in color from coal 
dust. Microscopically "morning sputum" consists of mucus, in which 
may be detected viscid streaks arising from the goblet-cells and more 
watery portions from glands. Embedded in this sputum are rows of cells, 
both epithelial and pus, loaded with coal pigment and myelin. In addition 
are non-nucleated cell-like masses (probably degenerated epithelial cells) 
and pus-cells clumped together in balls. These latter as a rule contain 
no pigment. 

Sputum is raised from the trachea by coughing unless there is so much 
that it actually flows from the mouth. But there are some patients who, 
although they should furnish sufficient sputum for examination, persist 
in swallowing it. Most of these are children, or persons of filthy habits, 
or partially unconscious patients. The doctor is often rewarded for the 
time he spends urging such patients to expectorate. 

One of Doctor Osier's assistants created amusement by sitting persistently at the 
bedside of a case with pneumonia and begging her to expectorate. At last he obtained 
a very little sputum, but it contained tubercle bacilli, and the hospital record for the 
early diagnosis of the pneumonic type of pulmonary tuberculosis was broken. 

Among the ways to get the swallowed sputum are, by washing out the 
stomach, and, in children, by examining the stools. Findlay recommends 
that in the case of young children the examiner cover his finger with gauze 
and with it tickle the child's pharynx to stimulate coughing. As the 
sputum rises in the throat it can be caught on the gauze. 

The patient must be carefully taught to avoid expectorating into the 
specimen cup saliva, nasal and pharyngeal mucus, food, etc. 

Amount. — A record of the amount of sputum expectorated each 24 
hours is sometimes of aid in following the progress of a disease. In some 
rare cases, although the cough is severe, the bronchial secretion is so scanty 
and so viscid that practically none is expectorated. This occurs in "dry" 
bronchitis, often in diffuse bronchitis, in incipient tuberculosis, in post- 
influenzal bronchopneumonia, rarely in acute lobar pneumonia and caseous 
pneumonia. Large quantities of sputum are expectorated in some cases of 
chronic bronchitis, of advanced tuberculosis with large cavities, of bron- 
chiectasis, of pulmonary gangrene, while in cases of edema of the lung, 
of albuminous sputum following thoracentesis, of pulmonary hemorrhage, 
of perforating pleural exudate and of rupturing lung abscess the serum, 
blood or pus evacuated through the bronchial tree may literally pour 
from the mouth and may even drown the patient. 



THE SPUTUM: GENERAL FEATURES 3 

The clinical chemist engaged in metabolism experiments must remember 
to take an abundant sputum into account since this may contain even 5% 
of the total nitrogen eliminated. 

Consistency.— Generally speaking, the consistency of sputum varies 
inversely with its total amount and directly with the amount of mucin it 
contains. Early in an attack of true bronchial asthma, in acute bronchitis 
and in pertussis the sputum is scanty and may be very tenacious. When, 
on the contrary, it is abundant and contains little mucin and much water, 
as in edema of the lungs and in chronic bronchitis which has denuded the 
bronchial tree of its mucous membrane, the sputum is very watery. A 
marked exception to the above rule is the sputum of croupous pneumonia, 
which, though abundant, will not spill from the inverted cup (see 
page 61). 

Reaction. — Fresh sputum is alkaline in reaction, but that which has 
stood for some time in the cup, or which has stagnated in the body, is 
usually acid, the result of bacterial fermentation. 

Character. — Mucoid sputum is glairy, transparent, tenacious and be- 
comes cloudy on the addition of acetic acid (due to mucin) . Such sputum 
is seen in acute bronchitis, pertussis, and early asthma. 

A mucopurulent sputum consists of mucus which contains pus-cells 
in sufficient numbers to cloud it. There are two forms of mucopurulent 
sputum. The first consists of clear mucus containing streaks and dots of 
pus while in the second the mucus and pus are more intimately mixed. 
If in the latter case the pus is present in small amount it gives the mucus 
a faint white haze; if it is relatively more abundant, a yellow or yellowish- 
green tint; if very abundant, the sputum may resemble pure pus. 

Purulent sputum is said to differ from pure pus in that the former 
contains mucus and so is more tenacious. But a purulent sputum may 
contain practically no mucus at all, as in the condition known as broncho- 
blennorrhea, present when the bronchial tree has been practically denuded 
of its mucous membrane. On the other hand the sputum certainly con- 
sists of almost pure pus when an empyema of the pleura perforates through 
the lung, or an abscess of the lung ruptures into a bronchus, or an abscess 
of a neighboring organ discharges through the lung, trachea, esophagus, 
or nasal passages. 

Serous sputum is watery and colorless and because of its high percent- 
age of albumin foams easily. One meets with it in edema of the lung, in 
perforating serous pleurisy, and in rare cases following thoracentesis. 

Color. — Bloody sputum may consist of almost pure blood or may 
gain its name from a much less, even a very slight, blood content. Such 
sputum may be due to trauma of the chest, hemorrhagic infarction of the 
lung, pulmonary gangrene, acute lobar pneumonia, caseous pneumonia 
(early), pulmonary tuberculosis, tumors of the lung, intense chronic passive 
congestion, or to "weeping" aneurism. As a rule the blood is mixed with 



4 CLINICAL DIAGNOSIS 

mucus and air and hence appears frothy. It may escape from the capil- 
laries by diapedesis, in which case it signifies a severe inflammation of the 
mucosa, or it may pour from a ruptured vessel. 

Sputum containing the derivatives of hemoglobin may be of almost 
any color. Formerly it was taught that variations in the number of blood- 
cells explained this variety of colors, but Traube proved that unchanged 
erythrocytes can color the sputum only red. Blood-cells free in the alveoli, 
bronchi, or lung tissue, however, soon disintegrate and the various oxida- 
tion products of their hemoglobin give the sputum that wide range of 
color — the various shades of red, brown, green, orange, yellow and choco- 
late seen, for example, in a subcutaneous bruise. Intact blood-cells in 
greater or less number may be found in such sputum, but the majority are 
pale and swollen. The best known example of sputum colored by modified 
free hemoglobin is the rusty sputum of pneumonia, the rusty color of which 
is due to an unknown derivative of hemoglobin. Another good example 
is the somewhat similar sputum which follows a small hemorrhage into the 
lung tissue or into a pulmonary cavity, while still another is met with in 
chronic passive pulmonary congestion, especially that due to mitral valve 
disease, in which sputum the characteristic light brown streaks and dots 
are due to masses of alveolar epithelial cells loaded with amorphous granules 
of modified blood pigment (Hertzf ehlerzellen) . The sputum of patients in 
whose lungs are areas cf necrotic lung tissue permeated with blood, such 
as occur in pulmonary gangrene, lung abscess or infarction, may have a 
uniform, dirty brown color due to masses of hematoidin crystals. Similar 
crystals are sometimes found in the sputum of cases with chronic passive 
congestion of the lungs. While the bile-stained sputum of a jaundiced 
person usually has no significance, yet the bile-stained sputum of a person 
who is not jaundiced may have value as in case of a liver abscess perforating 
through the lung. Such sputum may contain bilirubin or biliverdin. 

A definitely green sputum always demands an explanation. The pure 
mucoid sputum of a jaundiced patient with bronchitis may have a fine 
grass-green color due to biliverdin. Sputum of just the same color is seen 
in some patients (not jaundiced) during the lysis of an ordinary croupous 
pneumonia, in cases of pneumonia ending in abscess and in subacute caseous 
pneumonia. (It is interesting that Traube, Gesam. Beitr., ii, p. 699, 
187 1, first called attention to green sputum in "pneumonia" and cited 
only cases of caseous pneumonia.) Grass-green sputum may occur also 
in chloroma of the lung; while, finally, certain chromogenic bacteria may 
give the sputum this same color. 

The sputum in the various pneumokonioses deserves particular 
mention. The most common of these is antkracosis or induration of the 
lungs due to inhaled coal-dust. While the best examples of this are seen 
in coal miners, yet lesser grades are very common among city residents. 
The sputum in this condition usually is dirty gray, sometimes quite black. 



THE SPUTUM: GENERAL FEATURES 5 

Many granules, escaping expectoration, penetrate the bronchial mucosa, 
while others are swallowed and later make their way to the interlobular 
pulmonary lymph-channels, which they render beautifully visible. It is 
stated that gianules deposited in the interlobular tissue are never expector- 
ated unless freed by a destructive pulmonary process, yet some coal miners 
without any symptom of tuberculosis have continued to expectorate a 
black sputum for years after changing their occupation (Osier). Siderosis 
is due to the long continued inhaling of metallic dusts and occurs among 
workers in iron, bronze, brass, etc. The best examples, however, are 
furnished by mirror polishers, whose sputum is red with ferric oxide. 
Those who inhale much mineral dust suffer from chalicosis, called also 
"stone-cutters' phthisis," "grinders' rot," etc. These patients have very 
contracted chests and for years may have frequently recurring non-tuber- 
culous pulmonary hemorrhages. Practically all, sooner or later, become 
tuberculous. They are susceptible to various other pulmonary infections, 
as gangrene, which frequently leads to pneumothorax. Their sputum 
contains much of the mineral dust they inhale. 

Those who work with dry dyes, as methylene blue, have deeply colored 
sputum; bakers expectorate doughy masses; cotton mill operatives expec- 
torate masses of cotton; while particles of tobacco and of colored foods, 
drinks, medicines, are often found in the sputum of patients and may 
deceive the unwary. 

Finally, chromogenic bacteria, such as B. virescens, B. pyocyaneus 
and many others, which produce "sputum cup ward infections" may 
materially change the appearance of sputum. 

Air is present in the sputum in bubbles of varying sizes depending on 
the diameter of the bronchi furnishing the sputum, and on the efforts 
required to expel it. Sputum from lung cavities and from the larger bronchi 
may contain no air and hence will sink in water. This "sputum fundum 
petens" was formerly considered conclusive proof of a lung cavity but is 
met with also in acute tracheitis. 

Layer Formation. — The tendency of a sputum to separate into layers 
(layer formation) may be of assistance in diagnosis. In certain conditions, 
especially bronchorrhea, bronchiectasis, putrid bronchitis, and gangrene 
of the lung, the sputum is abundant, and, if collected in a tall jar. will 
separate into three layers: an upper, of frothy mucus; a lower, of morpho- 
logical elements, i.e., pus, tissue shreds, detritus, etc.; and a middle of the 
pus-serum, usually a cloudy, watery fluid. Often from the under surface 
of the upper mucous layer a sufficient number of streamers to constitute a 
fourth layer, consisting of the same material as the sediment, hang down 
in the pus-serum. 

Odor. — Fresh sputum is usually almost odorless. Sputum allowed to 
stand and that which has stagnated in the body soon gain a very positive 
odor. In tuberculosis and bronchiectasis the odor is heavy, sweet and 



6 CLINICAL DIAGNOSIS 

penetrating; that of a perforating empyema is said to resemble old cheese; 
that of putrid bronchitis and of many cases of bronchiectasis is fetid ; and 
that of gangrene is generally the worst of all. In pulmonary tuberculosis 
the odor of a patient's breath may be fouler than that of his sputum after 
it is cold. Some have claimed to have diagnosed on this evidence small 
tuberculous cavities before the physical signs of ulceration had appeared. 

MACROSCOPIC CONSTITUENTS 

Small masses of pus-cells are common in the sputum, their size 
indicating roughly the size of the bronchi from which they come. 

Fragments of necrotic tissue may be found in the sputum of cases of 
abscess or gangrene of the lung. These are sometimes quite large, even 
2 cm. long; those in tuberculous sputum are, as a rule, small, almost at the 
limit of vision. The fragments from a pulmonary abscess are yellow in 
color since permeated by pus- cells, those from other conditions may be 
dark from changed blood or black from coal pigment. Their nature is 
determined by the presence of elastic tissue. The discovery of even the 
smallest fragment of elastic tissue is important, for it proves the presence 
of an ulcerating lesion of the lung. To find these fragments the sputum 
should be squeezed out between two glass plates and studied with a hand 
lens. They are most numerous in the nummular masses from tuberculous 
cavities (see page 58). Fragments of necrotic cartilage from tuberculous 
ulcers of the larynx, trachea, or bronchi are sometimes found in the sputum. 
In a case of typhoid fever both arytenoid cartilages were expectorated. 
The demonstration of tumor fragments in the sputum may lead to the 
correct diagnosis of a doubtful condition. 

Dittrich's plugs are short cylindrical casts of bronchi, some scarcely 
visible, others as large as a bean. The size of the majority ranges from 
that of a millet seed to that of a mustard seed. The small plugs are opaque, 
yellowish-white and the larger ones dirty gray in color. If crushed between 
the fingers they emit a horrible odor. Microscopically, they consist for 
the most part of zooglea of bacteria, fatty acid crystals, fat droplets and 
cell detritus. They may contain also pigment granules, fragmented red 
corpuscles and hematoidin crystals. Flagellated protozoa and a lepto- 
thrix not yet thoroughly studied, but which takes a fine blue stain with 
iodine solution, have been found in them. The fatty acid crystals may be 
long and curved, or short, fine needles. The plugs contain but few intact 
cells. In some, perhaps fresher than the others, a few leucocytes can be 
recognized. These plugs may be found in the sputum of any putrid bron- 
chial disease, especially in putrid bronchitis and bronchiectasis, in which 
latter disease they are especially large. How these are formed we do not 
know (Hoffmann) , but the presumption is that they come from the smaller 
bronchi which open into a diseased or dilated larger bronchus. Somewhat 
similar plugs, shaped like a beechnut, come from the crypts of the tonsils. 



THE SPUTUM: MACROSCOPIC CONSTITUENTS 



1 


l'\ ' 


• ^**^"NQ" ■ 




S§^ 


^'s.J 



Fig. i. — Curschmann's spiral, from the 
sputum of a case of asthma. X 200 



Curschmann's spirals (Fig. i) are perhaps the most beautiful structures 
met with in the sputum. They are found in practically every case of true 
bronchial asthma at some time during its course. They have been reported 
also in the sputum of rare cases of acute bronchitis, acute lobar pneumonia, 
chronic pulmonary tuberculosis and in 
certain rare, interesting cases in which 
were combined the features of bronchial 
asthma and fibrinous bronchitis. In the 
sputum of one such case were found 
small fibrinous bronchial casts, the tips of 
whose branches were directly continuous 
with the "central fibers of typical spirals. 
Curschmann considered the spirals the 
result of a bronchiolitis exudativa. 

Two forms of spiral may be described. The first is a spirally twisted 
strand of mucus enclosing many leucocytes, especially the coarsely granular 
cells, and Charcot-Leyden crystals. The more beautiful form consists of 
a tight skein of mucus, the "mantle," wound around a "central fiber." 
These spirals are from i to 2 cm. or more long and 1 mm. broad. They 
may be branched. In the mantle are clumps of coarsely granular leuco- 
cytes, pigmented epithelial cells, ciliated cells and Charcot-Leyden crystals. 
The central fiber, made up probably of transformed mucus, is a very refrac- 
tive, spirally twisted strand, uniform in diameter, with a smooth contour 
or saw-edge. Some central fibers are small, from 0.5 to i/jl in diameter; 
some medium-sized, 3^; while others are thick, from 3 to 18^ in diameter. 
Ruge, who studied sputum hardened and cut into sections, found all these 
fibers solid, none with the lumen which others had described. Not all 
central fibers are as conspicuous as this. Some spirals have but a trace 
of a central fiber, others none. Some fibers end in a thread, others give 

off many lateral threads which radiate to the 
mantle. These finer fibers are often branches of 
larger ones. Some of the larger central fibers are 
lamellated, while others seem to be bundles of 
parallel threads, spirally twisted. These central 
fibers are not mere optical phenomena. They 
may partially project from the mantle, or lie in 
the sputum quite separate from it, and then are 
often twisted spirally. When alone they have the 
same significance as complete spirals. In the sputum of some cases one 
finds perfect spirals; in others, central fibers, some free and some with 
very imperfect mantles (see Fig. 1), while in still others free central fibers 
only (Fig. .2). 

The origin of the spirals is not understood. The central fibers certainly are not 
casts of the smaller bronchi, since their diameter is usually but one-tenth as great. 




Fig. 2. — Free central fiber of a 

Curschmann's spiral, from the 

sputum of a case of asthma. 

X 200. 



. 



8 CLINICAL DIAGNOSIS 

Schmidt considered that the conditions necessary for the formation of spirals were well 
preserved bronchial epithelium and a tough mucous secretion. Pie thought that the 
central thread represented the most twisted and therefore the most compact part of 
the spiral. Hoffmann claimed that the smaller bronchi are themselves spirals, which 
straighten out each time the lung expands, and that therefore tough mucus forced 
through them would assume a spiral form. Others claim that the cilia of the bronchi 
create spiral currents along the bronchi; others, that a straight band of mucus moving 
along a bronchus is whipped into a spiral by the tangential motion of the cilia of another 
bronchus at the point where these two unite. Gerlach stated that the three conditions 
necessary for their production were a small amount of very viscid sputum, very forcible 
respiratory movements and clear bronchi, three conditions which are best complied 
with in asthma. He claimed that the mantle and the central fiber both are formed at 
the same place in the bronchus but the central fiber later and is merely an optical expres- 
sion for that part of the mucous mass which has been twisted the most. We deny this 
emphatically. The central fibers are separate structures which may be found free of the 
spiral ; they are themselves twisted bands of fibers with markedly fewer revolutions per 
unit of length than the fibers of the mantle surrounding them. 

We have studied beautiful spirals about 
2.5 cm. long and 1 mm, broad, with central 
threads so refractive that they could be 
definitely seen with the naked eye. These 
central fibers or cores consisted of bundles of 
twisted fibers but the mantles surrounding 
them were much more tightly twisted. Some 
cores were bundles of fibers with very few 
turns indeed. It was evident on cross section 
that the mantles were composed of spirally 
wound sheets of mucus each enclosing var- 
3.— A spiral thread of mucus from the i OU s cells: squamous epithelial cells, very 
sputum. X 5- 1 1 11 j. • ■ 1 

many alveolar cells, some containing coal 

pigment and others modified hemoglobin, leucocytes especially eosinophiles, and cylin- 
drical cells, some with the cilia indistinctly seen and some of them goblet-cells. It was 
of interest that these cells were not mingled, but each variety occurred in groups or lines, 
large numbers in each group. The Charcot-Leyden crystals occur singly or in clumps. 
Some were quite large. One projected from a disintegrating eosinophile cell. In some 
fields full of leucocytes we searched in vain for one which was not coarsely granular. In 
one field was a strip of mucosa of cylindrical epithelium. Some spirals had no core while 
others consisted merely of a very large refractile central fiber- One spiral was partic- 
ularly interesting. It consisted of two strands of mucus, the inner thick, the outer 
thin, each rich in cells nearly all of which were coarsely granular leucocytes. These 
two strands, separate at one end, were twisted, the one within the other, into a spiral. 
For some little distance in the spiral each strand could be traced separately but the 
coil became tighter and tighter until they could no longer be distinguished. If the 
spirals are allowed to dry slightly the central thread becomes much more distinct. 

The sputum of another typical case of asthma contained spirals, immense numbers 
of eosinophile cells and a large number of epithelial cells, some of which were alveolar 
cells containing the various forms of pigment, while others were cylindrical cells, both 
ciliated and goblet in type. 

In another case were many central fibers, some without mantles and others with a 
few fibers of a mantle wound around them. The fibers forming these imperfect mantles 
were remarkably thread-like and of quite uniform diameter. Many of the coarse strands 
of mucus found in the sputum of bronchitis are spirally twisted (see Fig. 3). 




THE SPUTUM: MACROSCOPIC CONSTITUENTS 9 

Fibrinous Structures. — Under the title "fibrinous structures" we 
include all formations popularly supposed to consist of fibrin, although 
some contain none. In diphtheria one sometimes finds masses of whitish 
membrane in the sputum, which, if they come from the throat, larynx, 
or trachea, have no distinctive shape, but if from the bronchi they may be 
arborescent casts. In the sputum of pneumonia casts of the bronchial tree 
are often found, most of them small, but some very large (see page 63, 
and Fig. 2 5). These are brownish or reddish in color and contain blood 
and many leucocytes. Acute fibrinous bronchitis with similar casts in the 
sputum may accompany various fevers, such as typhoid fever, erysipelas, 
measles, smallpox, scarlet fever and acute articular rheumatism. It may 
also accompany exophthalmic goiter, pulmonary tuberculosis and mitral 
disease. Similar casts have been found in the albuminous expectoration 
after thoracentesis and after the inhalation of irritating vapors and gases. 
But the most interesting cases are those of chronic idiopathic fibrinous 
bronchitis, which condition has been well reviewed by Bettmann l (see 
page 73) . In the sputum of these cases, together with well formed arbores- 
cent casts and sometimes alone, one finds also unformed masses of the same 
material, evidently also from the bronchi. 

Whether any of the material in many of these structures is really fibrin 
or not is very doubtful. The tests for fibrin usually used are : the physical 
properties of the mass (its color, toughness, etc.), its tendency to swell 
and clear on the addition of acetic acid (which precipitates mucin) and 
the rapid effervescence on the addition of hydrogen peroxide. Hirsch- 
kowitz found, however, that one cast expectorated by a case of tuberculosis 
did consist of pure fibrin. 

Casts made up of the mycelium of fungi may be expectorated. In 
Osier's case a small cast consisted of a mass of aspergillus mycelial threads. 
Similar casts were expectorated for years by the case reported by Devilliers 
and Renon. 

Lung Stones. — Almost any mass in the sputum which is hard enough 
to wound the bronchial mucosa when it is expelled is called a "stone," 
whether cartilaginous, caseous, or calcareous in nature. The term should, 
however, be limited to masses of tissue or of inspissated exudate impreg- 
nated with lime salts which, dissected free by pyogenic infection, become 
foreign bodies in the air passages. 

Enchondromata and osteomata of the bronchi and lungs may be 
demonstrated in situ at autopsy, but among Poulalion's cases 2 we could 
find mention of none in which they were found in the sputum. At autopsy 
we find also pulmonary infarcts, areas of bronchopneumonia, miliary 
abscesses, the pseudotubercles of actinomycosis, cladothrix, or moulds, 
cyst walls, cyst contents and tumors which have become calcified, yet we 

1 American Jour. Med. Sci., Feby., 1902. 

2 Thesis, Pans, 1891. 



10 CLINICAL DIAGNOSIS 

know of no case in which a patient has expectorated calcified fragments 
of such tissues. Among the calcified masses which have been expectorated 
are fragments of calcified bronchial cartilage (Fraenkel) and of a calcified 
blood-clot (Hoffmann). But the vast - majority of lung stones found in 
the sputum are masses of calcified tuberculous tissue. These have been 
classified in two groups, bronchioliths and pneumoliths. 

Bronchiolites are formed by the deposit of lime salts in -the stagnated 
contents of a bronchus or bronchiectatic cavity. Some of these concretions 
have as nucleus a foreign body, for example, a cherry-stone or a grain of 
wheat. A few are arborescent; most of them are irregular and jagged and 
vary in size from that of a millet seed to that of a bean. They may be 
chalky or stony hard. Some "resembling coral, finely ramified and very 
hard" have been described. In one case the stone weighed 0.47 gm. and 
had 10 or 12 branches. In Atlee's case 3 the stone was % of an inch long 
and }i of an inch wide at the larger end. 

Pneumoliths may be fragments of calcified caseous areas of lung or 
masses of the calcified contents of a closed tuberculous cavity, or, and these 
are most numerous, fragments of calcified tuberculous bronchial lymph- 
glands. These masses are treated as foreign bodies and, with the aid of 
secondary pyogenic infections, ulcerate into a bronchus. In some of the 
first the structure of the lung tissue itself, even a few cell nuclei and tubercle 
bacilli, may be seen in sections of the decalcified mass. There are two dis- 
tinct, varieties of pneumoliths: the cretaceous, which are chalky in consist- 
ency, and the calcareous, which as a rule are small, hard and have a rough, 
rounded surface. Their size varies much. Some are as small as a millet 
seed while others the size of a pigeon's egg cannot be passed by the bronchus 
until they have disintegrated into fragments. 

Chemically, lung stones, whether pneumoliths or bronchioliths, contain 
for the most part calcium and magnesium combined with carbonic, phos- 
phoric, and sulphuric acids, with traces of ferric oxide and other metals. 
In some several minerals may be demonstrated, while others seem composed 
of but one calcium salt. It is seldom that one can say that a stone is a 
bronchiolith unless it is branched, or a pneumolith unless tissue structure 
can be demonstrated. Patients usually expectorate but one stone, seldom 
2 or 3, but some have expectorated very many, even 200 and in one case 
500. Poulalion suspects that all these were fragments of one large single 
concretion, while Hoffmann and others assumed in such cases a constitu- 
tional abnormality, an increased elimination of lime salts through the lungs, 
and named this condition " pseud ophthisis calculosa." Repeated hemor- 
rhages, "hemoptysis calculosa," often accompany the "bronchial colic" 
when the stone is expelled and are due to trauma of the mucosa. As a 
rule these hemorrhages are scanty, but some are severe. The presence of 

3 Stern, Deutsch. med. Wchschr., 1904, No. 39; Carlvon, Brit. Med. Jour., 1890, ii, 
P- 1474- 



THE SPUTUM: MICROSCOPIC CONSTITUENTS 11 

these stones may lead also to pulmonary abscess, pulmonary gangrene, 
or pneumothorax. 

The largest concretion I have seen was expectorated by a medical confrere after a 
period of eight months of ill health which at first suggested tuberculosis, then "asthma" 
and then for several weeks, pulmonary abscesses. This measured % inch in length and 
weighed 14 grains. His improvement following this illness was surprising. 

One of my patients, a young woman 38 years old, suspected of tuberculosis, expector- 
ated 22 such stones, irregular in shape, the largest weighing 2% grains and all together 
weighing 13% grains. The illness which precedes the expectoration of these stones is 
doubtless due to the secondary infection which sets the stones free. 

Among foreign bodies sometimes found in the sputum may be men- 
tioned teeth, cherry-stones, and coins (see also page 78). 

Fragments of the wall of echinococcus cysts or the daughter cysts 
themselves may be expectorated. 

MICROSCOPIC CONSTITUENTS 

The microscopical examination of fresh sputum though easy and often 
valuable is much neglected. A little sputum is spread upon a plate the 
base of which is half black and half white. The interesting particles are 
picked up on a hatpin and squeezed between a cover-glass and slide. It 
is important that the observer recognize at a glance the extraneous struc- 
tures, among which may be mentioned particles of food, particularly 
crumbs of bread, pieces of orange pulp or other fruits, drops of milk, bits 
of jams and preserves, fruit skins, tobacco, particles of meat containing 
elastic tissue which may lead to error in diagnosis and fragments of vege- 
table leaves. It is important to recognize also various threads, particularly 
fibers of vegetable tissue, linen, cotton, wool and silk (see Fig. 4). 

Cells. — The pus-cells of the sputum are usually polymorphonuclear 
finely granular leucocytes. As seen in the fresh sputum they are spherical 
and from 7 to 10/x in diameter. The majority are degenerated and contain 
fat globules, pigment granules, or glycogen granules. In asthma and in a 
form of bronchitis which has long been known as "eosinophilic bronchitis" 
the coarsely granular cells may greatly predominate. While their presence 
in great numbers suggests bronchial asthma it is of little value in diagnosis 
and on the other hand the absence of these cells certainly is not in favor 
of tuberculosis. 4 

To demonstrate these cells the sputum is spread on a slide, dried in the air and fixed 
over the flame. While still warm the slide is immersed for 5 minutes or longer in a 
°-5% alcoholic solution of eosin, washed in water and then counterstained for two 
minutes with a concentrated aqueous solution of methylene blue. 

The various epithelial cells found in the sputum demand careful 
study. Pavement cells may have come from the mouth, the pharynx, and 
the respiratory tract as low as the vocal cords. It is a valuable lesson for 

4 Hilderbrandt, Munch, med. Wchr., 1904, vol. 3 



12 CLINICAL DIAGNOSIS 

the student to scrape from the surface of the tongue a little epithelium 
and study the masses of these cells from the papillas among which are 
imbedded large zooglea of bacteria. Ciliated cylindrical epithelium, 
together with many goblet-cells, may come from the nose, trachea, or the 
bronchial tree. Cells with their cilia still intact are seldom seen in the 
sputum except in cases of asthma, of acute ulcerative processes and of 
very acute bronchitis. They soon lose their cilia. In very acute cases of 
asthma the sputum may contain clumps and even rather large sheets of 
cylindrical epithelium with the cells still ciliated. 

Tne alveolar epithelial cells present an interesting study. They are 
found in considerable numbers in nearly every sputum examined whether 
normal or pathological. They assume a large variety of forms and are 
often difficult to recognize. Until recently their origin was in doubt (see 



^ 



H 









, 



IV 



Fig. 4. — Extraneous matter common in the sputum. Threads of, A, linen; B, silk; C, 
cotton; D, wool; E, starch granules; F, guard cells from a lettuce leaf; G, squamous epithe- 
lium from tongue, with bacteria attached; H, tobacco, showing the surface of the leaf, the 
large cells stored with oil, and a spine from the surface. X 200. 

Hoffmann, Nothnagel's System, "Die Krankheiten der Bronchien"). In 
general these cells are from 4 to 5 times the size of a leucocyte, are oval, their 
protoplasm coarsely granular, and with one or several large, oval, vesicular 
nuclei. They are numerous in the sputum of cases of bronchitis which 
would indicate that in this condition the alveoli are involved as well as 
the bronchi. They occur most abundantly in the sputum of patients with 
inflammatory processes involving the alveoli, especially tuberculosis. In 
some cases of tuberculosis, however, these cells may fill the alveoli and yet 
none be found in the sputum. They are said to be ameboid on the warm 
stage; they certainly are active phagocytes. Some of these cells contain 
coal pigment (Fig. 5, a), that is, black granules, all of which are supposed, 
perhaps without sufficient proof, to be particles of carbon. The origin and 
composition of these granules were long disputed, but finally in one cell a 
single black granule was found which was unquestionably a particle of 
charcoal. It is these cells laden with inhaled dust which gives to the morn- 



RsSGRS 



5. 



„* 






m 


pi 


y 


- 


,%y h 










fi fll 


i 








^C 


















! 
j 




h 




i 


^ 1 
1 










\ 


y ^ 











Fig. 5.— Cells in the sputum, a, alveolar epithelium cells containing coal-dust; b, squam- 
ous epithelium cell; d, cylindrical epithelial cell; e and/, Herzfehlerzellen; g, cells snowing a 
peculiar degeneration; h, those with myelin droplets; i, one full of fat droplets; j, free myelin; 
k, red blood-cells; /, bacteria; m, free blood pigment. X 400. 



THE SPUTUM: MICROSCOPIC CONSTITUENTS 13 

ing sputum its dirty gray color. When very abundant the sputum con- 
taining them is smoky or dirty green. Some pulmonary conditions formerly 
bore the name "phthisis melanotica" because of the abundance of black 
pigment in the sputum. The fat globules (Fig. 5, i) which some of these 
cells contain are spherical, very refractile and glistening. Other alveolar 
cells contain one or more myelin globules which are irregular in shape, often 
concentrically marked, only slightly refractile, and have a dull greenish 
or bluish tint. These globules may be very minute in size, others are so 
large that one of them practically fills the cell (Fig. 5, h). Myelin is said 
by some to be a product of the degeneration of the protoplasm of the cells 
containing it, but others consider it a normal secretion of the bronchial 
mucosa, droplets of which these phagocytic alveolar epithelial cells ingest. 
Cells containing myelin are often numerous in the morning sputum of 
healthy persons, are very abundant in bronchitis and influenza, while the 
cases of " desquamatory catarrhal pneumonia" get their name from the 
sputum which resembles boiled sago because of the masses of alveolar cells 
filled with myelin droplets. Free myelin is sometimes present in such 
large amounts that it even exceeds the mucus in quantity, although in 
general the excretion of these two bodies runs approximately parallel. 
Free myelin is present in pale, non-refractive drops which vary much in 
size and more in shape (Fig. 5, /); some are concentric spheres, others 
club-shaped masses. The number and the size of these drops increase as 
the sputum stands. They resemble in appearance the myelin drops of 
nerve tissue. Similarly appearing and like named droplets are found in the 
urine and stools. The use of the name "myelin" in all these cases does 
not imply a chemical identity but merely a superficial resemblance. Small 
droplets of certain oils, of fatty acids, and of various neutral fats have the 
same appearance as these myelin droplets (Liebreich) . The myelin of the 
sputum consists chiefly of protagon, cholesterol and lecithin. It swells a 
little in water, is not destroyed at ioo° C, is stained yellow by iodine but 
poorly by aniline dyes, is not blackened by osmic acid, is easily soluble in 
alcohol and slightly soluble in ether and in chloroform. The alveolar cells 
which contain derivatives of hemoglobin are of particular interest. Modified 
hemoglobin may be present in amorphous granules, in scales of a brownish 
color, or as hematoidin crystals. "Hertzfehlerzellen" (see Fig. 5,/) is the 
name given to alveolar epithelial cells filled with golden yellow granules 
of a derivative of hemoglobin, provided these cells are found in large 
numbers and over a long period of time ; only then have they any diagnostic 
importance. These granules, which vary much in size, are not opaque or 
deeply colored but rather translucent. Certain cells seem to be diffusely 
stained by this substance. In chronic passive congestion, especially that 
due to mitral disease, these cells may be numerous enough to give the 
sputum a uniform rusty color, but more commonly they are clustered in 
masses which form dots and streaks of a reddish-brown color in the mucus. 



14 



CLINICAL DIAGNOSIS 



These cells may be found in the sputum in any condition in which red 
blood-cells escape into the alveoli. They are therefore numerous in chronic 
pulmonary congestion, pneumonia, infarction of the lung and after pul- 
monary hemorrhage. 

The red blood-cells (see Fig. 5, k) found free in the sputum are often 
well preserved, many are pressed or drawn out into long threads, but 
more are represented by masses of amorphous hemoglobin and of further 
modified pigment. The intact erythrocytes often are crowded into 
lines and masses in the mucus and are recognizable only by their color. 




Fig. 6. — Elastic tissue from lung. X 400. 




Fig. 7.- 



-Elastic tissue from lung showing alveolar arrangement. 
X 50. 



In judging the significance of blood in the sputum one should always 
bear in mind its quite numerous possible sources; the nose, the mouth, 
gums and the pharynx, as well as the bronchi and alveoli. 

Elastic tissue (Figs. 6 and 7) is one of the most important constituents 
found in the sputum. Before the discovery of Bacillus tuberculosis its 
presence was the best laboratory evidence of pulmonary tuberculosis, 
although it can also be found in the sputum of cases of pulmonary abscess 
or gangrene or cancer. It is our best proof of cavity formation due to 
any destructive disease. In some cases of tuberculosis elastic tissue is 
found before the tubercle bacilli. The masses of fibers of elastic tissue 
are often large enough to be visible to the unaided eye. Such is the case 



THE SPUTUM: MICROSCOPIC CONSTITUENTS 



15 



in pulmonary abscess and gangrene. In tuberculosis, in which molecular 
disintegration is the rule, these masses are very small and the most of 
this tissue is present as single fibers. To find it a little of the sputum 
is poured on a glass plate about 14 inches square and then pressed out 
by another glass plate about 6 inches square. The larger plate should 
rest on a dark background. The particles containing elastic tissue appear 
as small grayish-yellow dots which are easiest found with 'a small hand 
lens. Any suspicious particle may be exposed by sliding the upper glass 
away, picked up with a needle, crushed on a slide under a cover-glass 
and examined. It is in these masses also that one would have the best 







\ ■■■■■■Hi y£: J Beofajr 

Fig. 8. — Fatty acid crystals resembling elastic tissue in the sputum 
of a case of bronchiectasis. X 400. 

chance of finding tubercle bacilli. Instead of the glass plates for this 
examination some prefer to use Petri dishes, or wooden sputum boxes 
painted black, or crockery dishes with the base half black and half white. 
One must be careful not only to burn or sterilize these utensils after use 
for safety's sake, but also to cleanse the crockery and glass well in 
chemicals which will destroy all organic matter (a saturated solution of 
potassium bichromate in concentrated sulphuric acid is recommended), 
else there is danger later of finding the tubercle bacilli of a previous exami- 
nation. Particles of food will confuse the beginner. 

When no fragments containing elastic tissue are found, search should 
be made under high magnification for single fibers. For this it is well to 
select the grayish masses (presumably from a tuberculous cavity), or the 



16 CLINICAL DIAGNOSIS 

grass-green or slightly rusty particles (such as are found in subacute 
caseous pneumonia). Unless the cover-glass is well pressed down to give 
a very thin preparation the single fibers may be overlooked. 

To demonstrate elastic tissue some prefer to destroy all other organic matter. 
Mix 10 c.c. of sputum with an equal amount of 5 to 10% KOH or NaOH and boil in 
a porcelain dish until the mass is homogeneous. About 4 volumes of water are then 
added and the fluid is shaken up and centrifugalized. The value of this method is 
doubtful since the elastic tissue fibers will have lost their characteristic appearance 
and appear as pale, swollen threads. 

Elastic tissue stains, e.g., Weigert's stain, may be used to demonstrate 
this tissue in the sputum. Lord recommends the following method: 
From 0.5 c.c. to 1 c.c. of the thick purulent sputum is placed in a small 
Erlenmeyer flask and diluted with 15 to 20 volumes of distilled water. 

From 3 to 5 drops of KOH are added 
(as small an amount of KOH as possible 
should be used in order to keep the spe- 
cific gravity low) and the mixture gently 
warmed over the Bunsen flame until the 
sputum is dissolved. The solution is then 
sediment ed by means of the centrifuge, the 
supernatant fluid decanted and smears 
made from the small amount of sediment 
on cover-glasses by means of the platinum 
loop. These are dried in the air or over the 
Bunsen flame, fixed b}^ heat and covered 
with Weigert's elastic tissue stain. A con- 
venient method is to slip the cover-glass 
into a large test-tube containing enough 
fig. 9— a leptothrix form in the sputum, of the stain to cover it, and to immerse 

resembling elastic tissue. X 400. ,. . • . ,.. , M . « 1 ,1 

this in the boiling water of a water bath 
(a free flame cannot be used since the alcohol will ignite) for about 5 
minutes. The preparation is decolorized in alcohol (95%), dehydrated 
with absolute alcohol, cleared with xylol and mounted in balsam. The 
elastic fibers appear dark blue or almost black. 

The fibers of elastic tissue in fresh sputum (see Fig. 6) even when single 
usually may be recognized. In a thin specimen they stand out as very 
distinct, coarse, sharp, blackish fibers, characterized by their intense 
refr activity, their wavy outline, sharp edges, an uniform diameter and 
curling ends. They often branch. They are insoluble in ether and in 
potassium hydroxide. They are unchanged on warming. Pressure does 
not produce in them any varicosities. One should not confuse them with 
fibrous tissue, fatty acid crystals (see Fig. 8), bacteria, or 'vegetable fibers. 
The fibers of fibrous tissue appear in bundles of fine, wavy threads which 
have not the coarse, black, refractive appearance of elastic tissue. (For 




THE SPUTUM: MICROSCOPIC CONSTITUENTS 



17 



the fatty acid crystals see page 15.) The chains of bacteria are very 
confusing, especially certain leptothrix forms (see Fig. 9). These may be 
found in the fresh sputum, but more often in that which has stood for 
some time. They have not the same diameter, refractility or wavy out- 
line as elastic tissue fibers and yet the long chains of these organisms 
sometimes are arranged in a beautiful network which may resemble closely 
the framework of ah alveolus. Under the high power, however, one can 
make out the difference. Vegetable cells and fibers are as a rule much 
coarser, are irregular in outline and quite different in appearance. The 
elastic fibers from the muscle tissue of the food are of the same tissue but 




Fig. 10. — Elastic tissue from saliva; origin, the food. X 400. 



as a rule are coarser and more irregular in outline (see Fig. 10). To exclude 
these the teeth should be well brushed before the specimen is collected. 
Mould mycelial threads should give no confusion (see page 45). 

The presence in the sputum of elastic tissue, apart from that from the 
food, is proof of disintegration of the lung and, in about 90% of the cases, 
evidence of tuberculosis. It is found also in gangrene, abscess and neoplasm 
of the lung. 

The fibers of elastic tissue from the lung when in groups may present 
three arrangements which often suggest their source : fibers from the alveo- 
lar walls are long and branching and may preserve the outline of one or 
several alveoli (see Fig. 7) ; fibers from the bronchial walls are long and 
2 



18 CLINICAL DIAGNOSIS 

narrow, often fragmented and occur singly or in small groups of two or 
three clustered closely together in an elongated network; fibers from the 
walls of arteries appear as sheeting of elastic tissue. 

Tissue fragments from ulcers of the larynx contain a coarse network 
of short, interwoven, elastic fibers. 

Crystals are present only in putrid sputum, especially that of putrid 
bronchitis, bronchiectasis, tuberculosis and gangrene. - Fatty acid crystals 
are found in clusters. When very long they ma}' resemble elastic tissue 
(see Fig. 8) or microorganisms, but they are usually relatively thick, with 
stiff curves and pointed ends. If pressure is made on them by tapping 
on the cover-glass varicosities will result. These crystals are soluble in 
potassium hydroxide and in ether. (The specimen should be dried before 
ether is added.) If the slide is warmed they will disappear and fat droplets 
will appear in their place. Cholesterol crystals (see Fig. 22) are rare in the 
sputum and occur usually in connection with fatty acid crystals. Leucin 
and tyrosin (see pages 253, 254) may be found in sputum which has de- 
composed in the air passages and in the contents of a pulmonary abscess. 
The sheaves of long, black, refractive needles of tyrosin are more easily 
recognized than are the spherules of leucin. To demonstrate the latter, 
it is usually necessary to evaporate the sputum. Triple phosphate crystals 
(see page 248) and calcium oxalate crystals (see page 250) also occur in 
putrid sputum. Hematoidin, in rhombs or in needles (see page 2 53) , is seen 
in the sputum in cases of abscess of the lung, of perforating empyema and 
of a liver abscess discharging through the lung, but seldom after pulmonary 
hemorrhage in which case the extracellular hemoglobin appears chiefly as 
amorphous granules. 

Char cot-Ley den crystals (see Fig. 74) are long, narrow, diamond-shaped 
crystals which resemble two very sharp pyramids with their bases together. 
They have a slight yellowish refr activity and seem quite brittle. They 
vary greatly in size and are found singly, in groups, or in clusters. They 
are soluble in hot water, in mineral acids, and in alkalies and stain red with 
eosin. Viewed in the direction of their long axis one can see that they are 
hexagonal on cross-section, therefore cannot be identical with Bottcher's 
spermin crystals, with which they formerly were confused. That these 
crystals are derived from coarsely granular leucocytes is suggested by the 
fact that they are found only where these cells are increased, as in the 
sputum in asthma, in the blood in myelogenous leukemia and in the stools 
of patients with intestinal parasites. That they are products of decomposi- 
tion is shown by the increase in their number in a specimen of sputum 
left for some time in a thermostat. 

PLANT PARASITES 

The bacteria, of which usually there are large numbers in the sputum, 
are chiefly saprophytes, some from the mouth and others added later by 



THE SPUTUM: PLANT PARASITES 19 

the cup or the air. These multiply very rapidly at room temperature. 
The chromo genie organisms are mentioned on page 5. 

To obtain from the sputum the organisms from the lower air passages 
the patient washes his mouth well, cleanses his teeth thoroughly and then 
expectorates into a sterile cup. The sputum is at once rinsed several 
times in sterile physiological salt solution and cultures made. Among these 
are many saprophytes which nourish in the bronchi in cases of chronic 
bronchitis, tuberculosis and especially in bronchiectatic cavities. These 
saprophytes explain the decomposition of stagnant sputum. One finds 
also pathogenic staphylococci and streptococci which may be primary or 
secondary invaders of the bronchi. In tuberculosis they aid and perhaps 
explain the destructive processes in the lungs and cause most of the compli- 
cations and sequelae of this disease. 

Micrococcus Aureus — Staphylococcus Pyogenes Aureus (Fig. 11). — This 
organism is a coccus which ordinarily appears in clusters, hence the name 
"staphylococcus," but when actively 
growing in tissues usually occur as 
diplococci. The individual organisms 
are spheres a little less than iju in 
diameter. It is easily stained in all 
aniline dyes and is not decolorized 
by Gram's method. It is not a flagel- 
lated coccus. It grows well on all 
ordinary media. If grown directly 
from the animal body its colonies will 

SOOner Or later develop a golden Vel- FlG - i^— Micrococcus aureus. Photomicrograph 
r o j by Dr. Thomas M. Wright. 

low pigment, seen first at the edges 

of the thick, glistening, dull white colonies and later spreading throughout 
the whole growth. The most pigment is produced on potato. This organism 
liquefies gelatin, produces acid in milk and ferments nearly all sugars with 
gas production. It is pathogenic to animals, producing either a fatal 
septicemia — or, if the animal survives for a few days, a pyemia with 
multiple abscesses. In man it is the common pus-producing organism, 
causing local foci, as boils, abscesses, etc. 

Staphylococcus pyogenes albus is also one of the important pyogenic 
organisms. It differs from Staphylococcus pyogenes aureus only in that 
it produces no yellow pigment. Many think that it is really a variant 
strain of aureus. 

Staphylococcus epidermis albus is a normal inhabitant of the deep layers 
of the skin. It is quite similar to Staphylococcus pyogenes albus but is 
less pathogenic and a feebler grower. 

The Streptococcus Group. — The single organisms of the chains 
of streptococci are cocci about i/z in diameter which grow in chains of 
from 2 to 100 or more, each slightly flattened against its neighbors. 




20 CLINICAL DIAGNOSIS 

These chains have no capsule as have the chain forms of Diplococcus 
lanceolatus and Streptococcus mucosus. They do not decolorize by Gram's 
method. They grow on all ordinary media, but are feeble growers, forming 
colonies so minute and translucent that they are easily overlooked and 
which are quickly overgrown in mixed cultures. In the search for this 
organism in the urine, sputum, pus, etc., bacterioscopy is often of more 
value than are cultural methods. There certainly are several varieties 
of streptococci and yet no classification is quite satisfactory. Some strains 
do not grow at room temperature; some coagulate milk, others do not; 
some produce a growth, i.e., a sediment, in liquid media with rather dis- 
tinctive physical properties; some grow on gelatin, which they do not 
liquefy, others do not; some hemolyze red blood-cells, others do not; some 
produce pigment; many differ in their ability to ferment carbohydrates, 
all are quickly killed off in acid-reacting media, all are insoluble in bile 

and all decolorize litmus milk. Cer- 
tain varieties are definite enough to 
deserve mention. 

Streptococcus pyogenes (Fig. 12), 
formerly almost a "group" name 
since the other varieties described 
at the same time were not long 
accepted, is an intensely pathogenic 
organism for man, producing rapidly 
spreading inflammations, e.g., ery- 

Fig. 12.— Streptococcus pyogenes. Photomicro- sipelas, with much necrosis but With 
graph by Dr. Thomas M. Wright. . . 

little pus production. It is especially 
prone to cause inflammation of the serous membranes. It is an important 
cause of septicemia and pyemia. It is the most widely spread of all 
pathogenic organisms. 

Schottmtiller, who classified these organisms on the basis of their 
growth on blood agar (in this technic at least 5% of defibrinated blood 
should be used and all readings made at the end of 24 hours incubation), 5 
designated this as Streptococcus longus pathogenies sen erysipelatos, the 
inciting agent in erysipelas, which produces in blood agar about the colony 
a wide, clear zone of hemolysis due to an active hemolysin and without 
methemoglobin formation. Blake would include all those organisms under 
the name Streptococcus hemolysans (or hemolyticus) which produce a true 
hemolysin. On the basis of complement fixation tests Kinsella and Swift 6 
decided that this variety is homogeneous, consisting of members which 
are nearly identical. Opie 7 found this organism in the mouths of 1 of 
each 4 or 5 healthy soldiers at Camp Funston. It seemed to play an 

5 Blake, Jour, of Med. Research, vol. xxxvi, Mch., 1917. 

6 Jour, of Exp. Med., Aug. 1, 1918, xxviii, p. 169. 

7 Jour. A. M. A., 1919, vol. 72, p. 108. 



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THE SPUTUM: PLANT PARASITES 21 

insignificant part in the production of pneumonia but a very serious part 
in the causations of complications. It evidently was important in bron- 
chitis. It would seem able to pass through a pneumonic lung and appear 
in the blood-stream or in the pleural cavity. Lucke, Wight and Kime 8 
found at Camps Taylor and Knox that this streptococcus was the organism 
of a third wave of the "flu" infection, the second "wave" or "crop" being 
due to pneumococci and non-hemolytic streptococci. Cecil 9 found that 
24.7% of the pneumonias of this epidemic were due to streptococci and 
of these 24% to streptococci of the hemolytic type. The mortality of the 
streptococcus cases was over 30% while the general mortality for pneu- 
monia was only 15%. Of the streptococcus cases 24.9% developed em- 
pyema while of the pneumococcus cases, but 12.9%. 

Streptococcus mitior seu viridans (Schottmuller) produces green colonies 
in blood agar and changes oxyhemoglobin to methemoglobin without the 
production of a true hemolysin. It is rarely hemolytic and then only 
apparently so since the red cells in the clear zone do not lose their structure ; 
but it may form a narrow, clear zone about its green colonies. The name 
Streptococcus alpha hemolysans has been given to these organisms. Krum- 
weide and Valentine 10 considered the viridans not one type but a hetero- 
geneous group. One of these types which does not produce hemolysis nor 
methemoglobin has been named Streptococcus saprophytics but is classified 
now as viridans. Blake includes under the term Streptococcus viridans 
all which transform oxyhemoglobin to methemoglobin and includes also 
all which produce no change in blood. In this group he recognizes 
Streptococcus buccalis which will ferment lactose but not mannite, Strepto- 
coccus Jecalis which mil ferment both and Streptococcus equinus which 
ferments neither. Streptococcus buccalis has been obtained from the urine 
in acute nephritis, from alveolar abscesses, from the heart valve in endo- 
carditis, from tonsils and mouth, from the sputum in pneumonia and in 
bronchitis and from the blood in septicemia; Streptococcus fecalis, from the 
tonsils, from alveolar abscesses, from the urine in acute nephritis and from 
abscess in pyemia ; while Streptococcus equinus, from the tonsil and from 
the heart valve in endocarditis. On the basis of fermentation reactions 
Kinsella and Swift 11 assigned the 28 strains they studied to the fecalis 
group, the mannite fermenters and the salivarius group. The results of their 
complement fixation tests do not, however, justify this grouping and 
show the group to be heterogeneous. Holman 12 has given a much more 
elaborate classification. 

Streptococcus mucosus (Schottmuller) is an organism so similar to Micro- 
coccus pneumonias that many doubt that it is a separate bacterium and 

8 Arch. Int. Med., 1919, vol. 24, p. 154. 

9 Jour. A. M. A., 1918, vol. 70, p. 728. 

10 Jour, of Infect. Diseases, 1916, vol. xix, p. 760. 

11 Jour. Exp. Med., June, 1917, xxv, p. 877. 

12 Jour. Med. Research, 1916, xxxiv, 177. 



22 CLINICAL DIAGNOSIS 

group it with this organism under the name Pneumococcus mucosus. It 
differs from the former 2 varieties also in that it is bile-soluble. The 
organism, named also Streptococcus mucosus capsulatus, was found by 
Buerger 13 in the cerebrospinal fluid of a fatal case of acute meningitis 
and in the mouths of 6 normal persons. It is held responsible for some of 
the epidemics of "influenza," or at least of some of the complications. It 
is sometimes highly virulent and in human blood it is nearly unsusceptible 
to phagocytosis. Obtained from the blood and secretions of inoculated 
mice, it is found to occur chiefly as round, biscuit-shaped, or slightly lancet- 
shaped, encapsulated cocci, most of them in pairs, but always a few in 
chains of 4 or even 6, often irregularly arranged. The capsule is wide 
and easily broken. When this organism grows in pairs it resembles Micro- 
coccus pneumoniae but the cocci are never as uniformly and definitely 
lancet-shaped and the capsules are wider and show no traces of the con- 
strictions partially separating the cocci so often seen in the capsules of the 
organism of pneumonia. Streptococcus mucosus capsulatus grows luxuri- 
antly on serum agar and glucose serum agar. On blood agar it produces 
a mucoid growth with a dark -green zone. This is a very important point 
in its identification. It remains encapsulated through many subcultures. 
The colonies have the same watery, almost transparent, appearance as have 
those of Micrococcus pneumoniae, but they grow faster and tend to coalesce, 
so that the surface of the slant medium is finally covered by the' growth. 

Micrococcus catarrhalis is a common inhabitant of the respiratory 
passages of persons who are healthy as well as of those who are diseased. 
It is often seen in the nasal secretion and sputum of patients with common 
colds, it may be the cause of mild catarrhal inflammations and has been 
carefully studied in connection with epidemics of influenza. When this 
organism occurs as biscuit-shaped diplococci, and especially when intra- 
cellular, as is so often the case, it closely resembles in morphology the 
Gonococcus and Meningococcus intracellularis. It grows easily, however, 
on all simple culture media, which the gonococcus does not. Some strains 
so resemble the meningococcus that special media are required to differen- 
tiate them. For this purpose Dunham proposed a medium of sheep serum 
containing 1% of glucose. On this medium the meningococcus produces 
acid in 24 hours but no coagulation, while Micrococcus catarrhalis produces 
either acid and coagulation or an alkaline reaction. When Micrococcus 
catarrhalis grows in chains it resembles Streptococcus pyogenes, except 
that the individual cocci are larger and can be decolorized by Gram's 
method. Most recent writers (e.g., Holt) consider this an organism 
unworthy of serious attention. It has never been isolated from the blood 
or known to cause a general pyemic infection (Hasting and Boehm). 

Bacillus Tuberculosis (Fig. 13). — Possibly on no single clinical test 
is as much human interest centered as on the search for Bac illus tuberculosis 

13 Mt. Sinai Hosp. Reports, 1907, Centralbt. f. Bakt. Orig., vol. 41, p. 314 



THE SPUTUM: PLANT PARASITES 23 

in the sputum. A few drops of sputum are dried and stained in a certain 
manner. If in this specimen the observer finds one rod of a certain color 
and shape he has in the past been willing on this evidence alone to insist 
on an entire reorganization of the patient's life and to condemn him to 
exile from home for months or even years. Fortunately within the past 
few years wiser counsel has prevailed and fewer doctors rest so momentous 
a question on the stained specimen alone. Some have made it a practice 
to send the sputum of each of their patients with a cough to the State 
laboratory and make their diagnosis from these reports. The laboratory 
man can help the physician make his diagnoses but is not in a position to 
make them for him. One should in each case interpret the laboratory 
report in terms of the clinical history and the physical examination. 

Among the errors inherent in this test are the following: First, the 
specimens may have become wrongly labelled. 

Second, a red bacillus in the stained 
specimen with almost correct morphol- 
ogy may not be Bacillus tuberculosis; 
it may be some other acid -fast organ- 
ism from the nose or mouth, from food, 
milk or water. 

Third, this bacillus may be Bacillus 
tuberculosis but not from the patient 
under consideration. The dust of the 

air Which settles in his Sputum CUp Fig.. 13.— Tubercle bacilli stained with carbol- 

^ x fuchsm and decolorized with nitric acid. X 900. 

often includes this germ; the sputum 

cup may not have been cleaned perfectly (it may have been sterilized 
properly, but that is another matter J and so we find in the sputum bacilli 
from some other patient. The same danger applies to the hatpin we use 
in spreading the specimen, to the glass plates on which we pour it, and 
especially to the slide and cover-glass on which the specimen is made. 

Fourth, the tubercle bacillus we find may have come from the sputum 
of our patient and yet he ma}/- not have tuberculosis ; he may have 
inhaled dust containing dead or living tubercle bacilli and these we 
find in his sputum. 

The above mistakes are to be feared in case one or a few bacilli are 
found on a single examination. Therefore it should be an invariable rule 
to confirm a positive report at least once, taking every precaution to rule 
out all chances of such errors (see page 59). 

On the other hand, a negative report may lead to error. For instance, 
in the sputum of from 60 to 75% of early cases of tuberculosis no tubercle 
bacilli can be found; or, we may not have selected for examination the 
proper portion of sputum; and, lastly, the bacilli may not take their specific 
stain. In case of doubt the organism in question should be grown on media 
or injected into a guinea-pig (see page 27). And yet so valuable is this 




24 CLINICAL DIAGNOSIS 

clinical laboratory test that notwithstanding all these possible errors the 
test is one of the most valuable we have. The routine of this examination 
is as follows: 

The patient should at night cleanse his mouth and teeth and the next 
morning expectorate the first sputum he can raise into a perfectly clean 
cup. The sputum is then spread on a glass plate and the caseous particles 
already mentioned should, if present, be selected for the "examination, 
but if none are found, smears should be made from the small bloody or 
purulent masses. The bacilli may be found in the masses of blood of the 
initial hemoptysis. In advanced cases, bacilli may occur in goodly numbers 
in practically all portions of the watery mucoid as well as of the muco- 
purulent sputum. 

In case but very few bacilli are present the sputum should be spread 
on a large glass, squeezed under a smaller glass and careful search made 
with a hand lens for proper particles for examination. This is the best 
method we have. Others prefer to render the sputum homogeneous and 
fluid and then either to precipitate the bacilli to the bottom of the tube 
or to salt them to the top. 

The best quick way of obtaining a sputum sufficiently homogeneous and fluid is 
Lofner's lA modification of Uhlenhuth's antiformin method. Antiformin is supposed to 
destroy all bacteria except those which are acid-fast and all other formed elements 
as well. A measured quantity of sputum (5, 10 or 20 c.c.) is mixed in a Jena flask with 
an equal quantity of 50% antiformin (which contains NaCIO and NaOH) and boiled 
not over 15 minutes. The solution will foam considerably and become somewhat 
brownish in color. To each 10 c.c. of the fluid are now added 1.5 c.c. of a mixture of 
chloroform (1 part) and alcohol (9 parts). The whole is shaken vigorously until a fine 
emulsion is produced and then some of this emulsion is centrifugalized for 15 minutes. 
The heavier constituents, including some of the bacilli, collect in a film just above the 
chloroform. The supernatant fluid is poured off, the film is transferred to a cover-glass, 
or better, directly to the glass slide, and the excess of fluid is absorbed by filter paper. 
A drop of egg albumin mixed with carbolic acid (enough to give a 0.5% solution), or, 
better still, some of the original sputum not treated with antiformin, is added to fix 
the sediment to the slide. This sediment thus prepared is spread by a second slide or a 
clean hatpin into a thin smear, allowed to dry in the air, fixed in the flame, and stained. 
No cover-glass is necessary. The cedar oil is put directly on the stained smear. One 
can spread the smear more easily by holding the slide at such distance above a flame 
that it is slightly warmed. 

Another method is that of Paterson, who adds 2.5 c.c. of antiformin to each 10 c.c. 
of sputum and as soon as the sputum is dissolved pours the resulting fluid into centrifuge 
tubes. These centrifugal tubes when not in use are kept in a jar of potassium bichromate 
and sulphuric acid, and are well rinsed with distilled water before using. The tubes are 
stoppered with new corks, shaken, and allowed to stand 24 hours at room temperature 
or from 4 to 6 hours in a thermostat at 30 C. The tubes are then again shaken and 
centrifugalized, the supernatant fluid poured off, the tubes refilled with sterile physio- 
logical salt solution, shaken and again centrifugalized. This is repeated a second time. 
The sediment is then transferred to a slide and treated as above. 

One attempting to centrifugalize bacteria suspended in a fluid should remember that 

14 Deut. med. Wchs., Oct. 27, 19 10, vol. xxxvi, No. 43, p. 1987. 



THE SPUTUM: PLANT PARASITES 25 

the reason why we so often succeed is that many of the organisms are attached to small 
masses of sediment which are heavy enough to be thrown to the point of the tube. The 
specific gravity of tubercle bacilli lies roughly between i.oio and 1.080, and if, as often 
happens, the specific gravity of the fluid is heavier than that of the bacilli (the specific 
gravity of sputum varies from 0.929 to 1.2242) centrifugalizing will send the most of 
the organisms to the top rather than to the bottom of the tube. In case they are sus- 
pended in a fluid not coagulable by alcohol one can prevent this by diluting the fluid 
with an equal volume of alcohol. The mixture is then so light that even the free bacilli 
will sink to the bottom. Others prefer to add to the fluid an equal volume of a 25% 
NaCl solution and allow the mixture to stand 24 hours. In this heavy liquid the bacilli 
will rise to the top. 

Staining. — The possibility of an almost specific stain for Bacillus tuber- 
culosis depends on the fact that this organism resists decolorization by 
acid and by alcohol. 

As a routine method we recommend the following technic which we 
shall describe first briefly, then in detail. The specimen, dried and fixed 
by heat on a slide, is covered with the Ziehl-Neelsen carbolfuchsin solution 
and heated over a small flame. Fresh stain is from time to time added to 
keep the specimen from drying and the glass from cracking. The staining 
solution covering the smear should actually boil for at least 1 minute. 
The stain is then poured off, the smear washed in water and dried with a 
blotter. It is then decolorized with acid alcohol until it will decolorize no 
more. Next, it is washed in water, dried with a blotter and covered, for 
about 5 seconds with Lofner's methylene blue. After the excess of this 
stain is washed off the specimen is again dried with a blotter and mounted. 
On examination the tubercle bacilli will be found stained red and all other 
organisms blue. 

The following points of technic deserve more detailed statement. Bacillus tubercu- 
losis is stained with difficulty but when once stained is also decolorized with difficulty. 
The entire specimen is therefore first overstained with a very penetrating dye which 
will certainly stain Bacillus tuberculosis. This will of course easily overstain all other 
bacteria. The specimen is then decolorized with a reagent which will remove the stain 
from practically all organisms except Bacillus tuberculosis, so that when cdunterstained 
with a blue or brown dye the tubercle bacillus alone will retain the original red stain. 

Ziehl-Neelsen' s carbolfuchsin mixture is the stain in common use for this purpose 
(Fuchsin 1 gm.; absolute alcohol 10 c.c; and 5% carbolic acid 10 c.c.) Bacillus tubercu- 
losis usually can be deeply stained in 1 minute in this solution at the boiling tempera- 
ture, or in 24 hours in the cold stain. By the rapid method the carbolfuchsin solution 
should actually boil for at least 1 minute, but 1 to 4 minutes are safer or the tubercle 
bacilli may not take the stain. Some prefer to submerge the slide or cover-glass 
in a flat dish full of the stain and to boil this over a free flame. Mere steaming is not 
sufficient. Even with the best of technic probably not all of the tubercle bacilli in 
the specimen will take any stain. 

The slow method is better and should be used if haste is not necessary since the 
specimens are much better stained. Heat certainly injures the specimens. The slides 
are placed vertically (to avoid a precipitate on the specimen) in a tall staining jar full 
of the carbolfuchsin solution and allowed to stand for 24 hours at room temperature 
(or in a thermostat). The further steps are those described above. 



26 CLINICAL DIAGNOSIS 

Having stained deeply as many of the bacilli in the specimen as possible, the next 
step is to decolorize all but the tubercle bacilli. The best reagent for this purpose is 
acid alcohol (2% HC1, some say 3%, in 80% alcohol). The slide may be dipped into a 
vessel of this acid alcohol, or this fluid may be repeatedly poured on and drained off 
the slide until the smear is almost colorless. A little time may be saved by gently warm- 
ing the slide during this process. Some decolorize under the low power of the microscope 
and are able to determine very accurately when the end point is reached. Another 
decolorizing fluid is 25% nitric acid, and still another is the very popular Gabbett's 
fluid which contains 25% sulphuric acid (see below). Gabbett's fluid, however, 
"burns" the specimen more than does nitric acid, and as a result the bacilli look too 
thick and are less distinctly beaded. Acid alcohol is far preferable to either sulphuric 
or nitric acids in aqueous solution since there is a large group of acid-fast organisms which 
may be confused with Bacillus tuberculosis but very few which are acid-alcohol-fast. 

After the specimen is decolorized it is washed in water and blotted with filter paper. 
If as a result any red tint returns, the acid is again applied. 

As counterstain Lofner's methylene blue (see page 38) is in general use. Lofner 
himself, however, used a 0.1% solution of malachite green (Malachitgriin crystallen 
Chlorzinkdoppelsalz, Hechst). The decolorized, washed and dried specimen is covered 
with this stain for about 5 seconds, the stain then washed off with water and the speci- 
men mounted. The very popular Gabbett's fluid, in which is not only the decolorizing agent 
but also the counterstain, contains enough methylene blue (1 to 2 gms.) to saturate 
100 c.c. of 25% sulphuric acid. By this method the specimen, after the excess of carbol- 
fuchsin has been washed off, is dried and then covered for 1 to 5 minutes with Gabbett's 
methylene blue. This is then washed off in water. If any pink tint remains, except in 
the thick portions, the smear is again covered with Gabbett's solution and again washed. 
This method is very popular since so easy. It is fairly satisfactory for the examination 
of those specimens in which tubercle bacilli are practically the only acid-fast bacilli 
to be found and especially if repeated examinations are to be made in cases the diagnosis 
of which is not doubtful in order to determine any increase or decrease in the number of 
bacilli present. 

Not all tubercle bacilli are acid-fast. This probably explains our 
failure to find them in the sputum of certain cases of proven tuberculosis. 
Since those tubercle bacilli which are not acid-fast will not decolorize 
by Gram's method, Much recommends that we search for Gram positive 
bacilli in the sputum in which we fail to find acid-fast organisms. 

It is also true that not all acid-fast rods seen in specimens are tubercle 
bacilli. Doubtless far too many specimens called positive should have 
been reported as negative. Some laboratory workers estimate that this 
error is present in about 10% of the positive reports made. 

Morphology. — The majority of tubercle bacilli found in the sputum 
are from 1.5 to 3.5^ long and about o.2ju wide. Some, however, are over 
ii/jl long, branch, are curved and resemble spirochete. Many, even of the 
shortest, are somewhat bent. Chains of these bacilli are sometimes seen. 

In stained specimens the bacilli are found scattered or in clumps and 
those in clumps may lie parallel or crossed. Beaded forms (from one to 
eight beads in each rod) resembling streptococci are common. These are 
considered degenerated forms. That the young bacilli stain less intensely 
than do the older forms is now disputed. 



THE SPUTUM: PLANT PARASITES 27 

Muck's granules, 1 ' minute acid-alcohol-fast granules, are considered to 
be fragments of tubercle bacilli. They are often found together with per- 
fect tubercle bacilli in specimens which have been treated with antiformin 
while in about 10% of the cases of tuberculosis one finds them and no 
perfect bacilli. 

Bacillus tuberculosis is not a spore-bearing organism and is as suscepti- 
ble to heat as are the other bacteria which do not produce spores; 16 i.e., 
all are killed in twenty minutes at 6o° C. 

Tubercle. bacilli will retain their virulence in dried sputum for from 3 
to 10 months. 

While other bacteria contain only from 1.7 to 10% of fatty matter, 
tubercle bacilli contain from 10 to 37%. This may explain their unusual 
staining characteristics. 

Animal Inoculation. — The theoretically best method of identifying 
tubercle bacilli is by inoculating guinea-pigs with the sputum under exami- 
nation. This never has been a popular method because Diplococcus 
lanceolatus is often present also and would kill the animal, and also because 
one must wait from 4 to 6 weeks for a result. These objections are not 
insuperable, however, for the sputum can be freed from the throat and 
mouth organisms by washing it (see page 19); also, since dead tubercle 
bacilli will produce tubercles, all organisms present may be killed by heat 
and then the sputum injected into a guinea-pig. To shorten the time one 
must wait for a result it was suggested by the work of Murphy and Ellis l7 
to expose the guinea-pig to massive doses of Rontgen rays which will 
inhibit the protecting action of the lymphatic tissue. 

This technic has been standardized by Eckford 1S as follows: The sputum is digested 
for 15 minutes with 4% potassium hydroxide after Petroff's method and is centri- 
fugalized and washed before injection. (If the material to be tested is not sputum but 
of a fibrinous nature it is digested in 15 9c antiformin until fluid. If it is a clear fluid 
it is merely centrifugalized.) 

From 0.5 to 1 c.c. of the material is injected both into the groin and into the abdo- 
men of the animal, which is then exposed every second day to a five-minute dose of X-ray 
for at least 3 exposures. (The dose is given with the Coolidge tube, the current 
60,000 volts, the spark gap 6 inches, the filter 0.5 mm. of aluminum and the target 
12 inches.) If a nodule appears and is still present in 2 weeks it is removed under a 
local anesthetic, smeared and examined. If a positive diagnosis cannot be made the 
animal is kept for at least 6 weeks and then examined. 

The Cultivation of Bacillus Tuberculosis. — Some of the best workers 
cultivate the organism for clinical diagnosis. 

Petroff s Medium}* — Petroff proposed for the clinical demonstration of 
Bacillus tuberculosis a medium which contains: 

15 Korber, Deut. med. "Wchschr., August 8, 1912. 

16 Rosenau, Bull, of Hygienic Lab., Sept., 1909, No. 57. 

17 Jour, of Exp. Med., 1914, xx, 397. 

18 Jour, of Lab. and Clin. Med., 1917, ii, 175. 

19 Jour, of Exp. Med., 1915, xxi, 38. 



28 CLINICAL DIAGNOSIS 

Egg (white and yolk), 2 parts by volume. 
Meat juice, 1 part by volume. 

Gentian violet, 1% alcoholic solution, enough to make a solution 
of 1 : 10,000. 

To make the meat juice, 500 gms. of beef or veal are infused in 500 c.c. of a 15% 
solution of glycerin in water. After standing for 24 hours the meat is- squeezed in a 
sterile meat press and the juices collected in a sterile beaker. 

The shells of the eggs used are sterilized by immersing them for 10 minutes in 70% 
alcohol or by pouring hot water over them. They are then broken into a sterile beaker 
well mixed and filtered through sterile gauze. 

About 3 c.c. of this medium are poured into each sterile tube and dried for 3 
successive days: on the first day at 85 C. until the medium is solid and on the second 
and third days for not more than 1 hour at 75 ° C. 

Under the most favorable conditions it takes at least 6 days for a single 
tubercle bacillus to grow to a visible colony. 

In sputum examination about 5 c.c. of fresh sputum and an equal 
amount of 3% NaOH are well shaken and left in the incubator for 20 or 
30 minutes until the sputum is fairly well digested. It is then made neutral 
to sterile litmus paper with 0.1N HC1, centrifugalized and the sediment 
inoculated into several test-tubes containing the above-described medium. 

Petroff obtained very uniformly satisfactory results. The cultures 
usually are pure since the NaOH and later the gentian violet should kill 
off all other bacteria. 

Bacillus tuberculosis grows better the longer it has been artificially 
cultivated in the laboratory. 

That this method cannot supplant the other methods is clear from the fact 
that a few strains have been found which fail to grow well on any medium. 20 

Bovine Tuberculosis. — It is now agreed that the bacillus of human 
tuberculosis and that of bovine tuberculosis are distinct types. Both are 
pathogenic for man, the bovine especially during infancy and early youth. 

Differentiation Between the Human and Bovine Types of Bacillus Tuberculosis.— 
The method described is that followed by Fraser 21 who employs tests described by 
Theobald Smith. In case the bacillus in question is to be isolated from diseased tissue 
this material itself is injected beneath the skin of guinea-pigs. In this way a pure culture 
of Bacillus tuberculosis may be obtained and the possible strains of saprophytic tubercle 
bacilli which may flourish as secondary invaders in lesions caused by more pathogenic 
organisms, and which grow on media more readily than the latter, are excluded. The 
inoculated guinea-pigs are permitted to live from 4 to 6 weeks, during which time 
careful records are kept of their condition. They are then killed and cultures from the 
tuberculous organs, especially the glands and spleen, are made by rubbing with a small 
platinum spud the diseased material onto the surface of the media. The tubes are 
sealed with paraffin. The media used are numerous. Theobald Smith 22 used dog's 
serum; Hiss and Zimmer recommend slants of agar to which rabbit's blood has been 
added, 1 to 2 c.c. to each tube; glycerin agar (containing 3 to 6% of glycerin) is also 

20 Corper, Am. Rev. of Tuberc, 1919, lii, 461. 

21 Jour. Exp. Med., October 1, 1912, vol. xvi, p. 432. 

22 Jour. Exp. Med., 1898, vol. iii, p. 451. 



THE SPUTUM: PLANT PARASITES 29 

used. In about 10 days a growth is usually apparent. A rapidly growing organism is 
likely to be human in type, while one which gives a weak and scanty growth is probably 
bovine. This test alone is not conclusive. 

If in the early and original culture the bacilli are long, slender, slightly curved and 
regular in shape and show granules, and if they tend in later cultures to grow in still 
longer forms the chances are that they are of the human type; if they are thick and short, 
from i to i.5yuin length, straight, not regular in shape, but some of them are spindles, 
others broader at one end, the bovine type is suggested. This test also is not conclusive. 

Glycerin inhibits the growth of the bovine, but stimulates that of the human, type. 
For this reason if a primary culture on glycerin medium grows luxuriantly from the 
beginning it is without much doubt the human type ; if sparsely or not at all, the bovine. 
(The medium used in this test is glycerin bouillon made of beef or veal with peptone, 
i%» glycerin, 6%, and rendered slightly alkaline, This is a very valuable test.) 

The "change of reaction" test proposed by Theobald Smith is almost conclusive. 
The main difficulty in this test lies in obtaining a suitable surface pellicular growth of 
the organism in question. To get this the organism is first cultivated on a tube of egg 
medium with a small quantity of glycerin bouillon in the bottom of the tube. The 
pellicle which extends over the surface of the bouillon is then transferred to an Erlen- 
meyer flask containing the test medium. This medium is glycerin bouillon originally 
neutral to litmus which must then be carefully titrated and brought up to standard 
acidity by adding 0.05 c.c. of 0.2N HC1. As the organism grows on this standardized 
medium and the pellicle develops, the acidity of the medium in the case of the human 
type increases, while in the case of the bovine type the acidity for a time diminishes and 
the medium may even become alkaline. To determine the change of reaction, 5 c.c. 
of the fluid are removed every 10 days, diluted to Ko its strength and titrated hot. 

The inoculation test is very valuable, perhaps absolute. If a rabbit weighing on an 
average 2000 gms. is inoculated intravenously with a known amount of the human 
bacillus (Fraser recommends 0.01 mg. of dried weighed pellicle emulsified and the emul- 
sion so diluted that 1 c.c. contains 0.01 mg.) the resulting lesions will be few and small 
and after a time show a tendency to undergo retrogression. Death as a rule occurs in 
6 months or longer and then often not from tuberculosis. If a bovine bacillus is used 
an acute disseminated rapidly fatal tuberculosis will develop. 

The bacillus of fish tuberculosis is similar in morphology to Bacillus 
tuberculosis and is rather acid-fast but grows at surprisingly low tempera- 
tures (15 to 30 C). 

The bacillus of avian tuberculosis closely resembles Bacillus tuberculosis in 
its morphology and staining characteristics. It grows on artificial media 
much more rapidly than does the human type and at a temperature from 
4i°to45° C, which is above that at which the latter will thrive (40 C). 
There is evidence that either of these organisms may by proper cultiva- 
tion and proper passage through animals be made to resemble the other. 

Acid-past Organisms. — The important group of acid-fast organisms 
deserves careful study. Supposed at first to be peculiar to but few bacteria, 
resistance to decolorization by acid is now known to characterize a large 
group, chiefly of non-parasitic organisms. Among these are 23 Bacillus 
tuberculosis, Bacillus leprae, the smegma group, the "milk and butter " 
bacilli (Rabinowitch), the timothy hay bacillus and the grass bacillus of 
Moeller, bacilli found in manure (Moeller), and a large group found in 

23 Borrel, Bull. Inst. Pasteur, May 30, 1904. 



30 CLINICAL DIAGNOSIS 

sewer water, in soil, etc. The majority of these organisms are of much 
greater interest to the hygienist than to the clinical microscopist, altogether 
the interest aroused by the report that it is easy to find Bacillus tubercu- 
losis in the circulating blood of consumptives (Rosenberger) demonstrated 
to the microscopist the necessity of examining his reagents, even distilled 
water, for acid-fast bacilli. 24 Several of these organisms have considerable 
clinical importance, as they are often mistaken for Bacillus tuberculosis. 
Chief of these are the smegma organisms which are found in those parts 
of the body where the secretions of the skin are allowed to collect, as around 
the genitals (where they were first discovered, whence their name, and 
where they abound) within the folds of the thighs and buttocks, in the 
axillae, in cerumen, etc. The smegma bacilli while acid-fast are not alcohol- 
fast and when grown in culture media many are not even acid-fast. It 
may be that the difficulty in decolorizing them by acids and the ease with 
which they are decolorized in acid alcohol are due to a fatty coating gained 
from the secretion of the skin where they were found and not to any 
quality of their own. Some strains of the smegma bacilli have a morphology 
identical with that of Bacillus tuberculosis, but in general they vary much 
more in size and appearance than does the latter. Acid-fast organisms, 
probably of the same group as smegma bacilli, are found also in the nose, 
in the coating of the tongue, in the tonsillar crypts, in the throat, in the 
stools, in the sputum, and in destructive diseases of the lungs, especially 
pulmonary gangrene. They are grown with difficulty, at first on media 
rich in human blood-serum or hydrocele fluid, but the later subcultures 
grow readily on glucose agar. 

These non-pathogenic acid-fast bacilli do not constitute a biological 
group. Indeed, their ability to withstand decolorization by acid is almost 
the only characteristic they have in common. While most of those we 
find in clinical examinations are shorter, thicker, and more homogeneously 
stained than Bacillus tuberculosis, yet some have exactly the same mor- 
phology. Practically all are decolorized by alcohol and most of them are 
not particularly acid-fast. It is to be borne in mind, however, that, as 
in the case of the smegma bacillus, the text book descriptions of these 
non-pathogenic acid-fast organisms are based on studies of the organisms 
as grown on artificial media, rather than as found in nature, while we study 
Bacillus tuberculosis not in cultures but as obtained directly from the host. 
Since the smegma bacilli grown on artificial media lose some or all of their 
acid-fast properties, it is possible that, should we obtain the other organisms 
in the same way in which we obtain the tubercle bacilli, i.e., immediately from 
the host, we might find them much more acid-fast than we now think and it 
is possible that they are a greater source of error than we realize. According 
to our present knowledge, however, they all are decolorized by alcohol and 
so this method should be our routine in searching for Bacillus tuberculosis. 

24 Burville-Holmes, Am. Jour, of Med. Sci., 1910, vol. xxxix, p. 99. 



THE SPUTUM: PLANT PARASITES 31 

Several 25 organisms of pseudo-tuberculosis which produce pathological 
lesions very similar to those of tuberculosis have recently been described, 
but none of these organisms themselves could be confused with Bacillus 
tuberculosis, so different is their morphology. 

Bacillus Leprae. — Bacillus lepras resembles Bacillus tuberculosis in 
that it is alcohol- and acid-fast. Morphologically it is similar although 
slightly more slender. Some claim that it can be cultivated, although 
with extreme difficulty, on media containing glycerin, and that the bacillus 
obtained from cultures is no longer acid-fast, but the chances are that no 
organism yet cultivated was Bacillus lepras. 

In cases of leprosy, Bacillus lepras is present in large numbers in the 
naso-pharyngeal, urethral, and vaginal secretions, in the saliva, and in the 
feces. Since the nasal lesions are among the earliest in leprosy and since 
it is easy to find this organism in the nasal secretion thus the early and 
satisfactory diagnosis of leprosy, is possible. Smears from these secretions 
are prepared and stained just as for tubercle bacilli, always using acid 
alcohol as the decolorizing agent. 

B. D., No. 3990, aged 59, admitted December 10, 1916, for destructive -lesions of 
fingers and toes for years supposed to be luetic, was stopped in the examining room 
by two internes who suspected it was leprosy and confirmed their suspicions before the 
visiting physician could arrive by smears of the nasal secretion, which showed remarkable 
numbers of Bacillus lepras. 

Streptothrix Pseudo-tube? xulosa {Streptothrix eppingeri). — This strep to- 
thrix was first found by Flexner 26 in the lungs of a patient with symp- 
toms suggesting pulmonary tuberculosis. Warthin and Olney 27 collected 
five such cases. While this streptothrix may be the primary invading 
organism it is oftener part of a mixed infection, as in cases of tuberculosis 
with cavity formation and of bronchiectasis. This organism grows in 
true branching threads which form large, entangled masses, even grossly 
visible as minutely grayish granules in a white, homogeneous, not bloody 
sputum. Some of the filaments are very long and thick, with short branches 
and without club-shaped ends. They are acid-fast, the carbolfuchsin 
giving them a beaded appearance, but they are slowly decolorized by 
95% alcohol and by 30% nitric acid. They often resist decolorization by 
Gabbett's stain. They stain by Gram's method. The streptothrix group 
is discussed by Claypole. 28 These organisms (Lord) usually grow readily 
but slowly on all culture media under aerobic conditions and at room as 
well as body temperature. Cultural peculiarities are variable. Under 
the microscope young colonies usually show the presence at the periphery 
of radially arranged and branching filaments. In smears from cultures 

25 Abbott and Gildersleeve, Centralbl. f. Bakt., 1902, xxxi, p. 547. 

26 Johns Hopkins Hosp. Bull., June, 1897. 

27 Amer. Jour. Med. Sci., 1904, vol. cxxviii. 

~ H Jour, of Exp. Med., Jan. 1, 1913, vol. xvii, p. 99. 



32 CLINICAL DIAGNOSIS 

rods with conical extremities may be observed as well as the slender branch- 
ing filaments. Bacillary and coccus-like forms may arise in fresh material 
as well as in older cultures by the breaking up of the filaments. A surface 
growth may be seen on bouillon or the formation of ball-like masses at 
the bottom of the tube without clouding of the medium. The results of 
animal experiments are inconstant. In some instances -local abscesses 
follow subcutaneous or intraperitoneal injection, while widely disseminated, 
yellowish, miliary tubercle-like nodules may follow intravenous inoculation. 
The nodules on histologic examination are infiltrated with leucocytes and 
have a more or less extensive central necrosis. Spore formation (chain 
sporulation) is thought to occur. 

The sputum in pulmonary streptothrix infection is more or less abun- 
dant, purulent and at times streaked with "blood. Cases with hemoptysis 
have been reported. In other cases the clinical features are those of 
empyema and, if the chest wall is perforated, suggest actinomycosis, while 
in still other cases the course resembles that of a pyemia. 

The leptothrix group of normal mouth organisms may flourish in abund- 
ance in the lungs, especially in putrid gangrenous disease. Their probable 
effect is to aid in the decomposition of the sputum. Miller has separated 
from the group formerly called " Leptothrix buccalis" several organisms 
among which are Leptothrix innominata, an organism which is unseg- 
mented, straight, but sometimes wavy, and from 0.5 to o.8ju broad. One 
always finds it in the tartar of teeth. This organism cannot be cultivated 
and is stained a pale yellow by iodine solutions. Bacillus buccalis maximus 
is an organism from 30 to 150^ long and 1 to 1.3^ broad arranged in long 
single threads or in bunches of parallel threads. It takes a deep blue stain 
with iodine. It cannot be cultivated. Leptothrix maximus buccalis is 
an organism somewhat longer than the last mentioned but otherwise 
similar except that it does not give the iodine reaction. For the mouth 
spirochetal, see page 41. 

Micrococcus pneumonia is a small, oval coccus, about i/z in longest 
diameter and usually arranged in pairs but often in chains. Even when in 
chains one can usually see that the individuals are oval, their long diameters 
in the line of the chain. The free ends of the diplococci are often pointed 
like a lancet (or better still, a candle flame), hence the former name, 
! ' Diplococcus lanceolatus. ' ' Micrococcus pneumoniae is a capsulated organ- 
ism whether in pairs or chains. There is always doubt about the identity 
of any non-capsulated diplococcus. 

Among the capsule stains the following are to be recommended. In 
all the important point is to avoid the use of pure water. 

Welch's Method. — The film, made from the fresh sputum on a glass 
slide, air-dried and then passed through a flame slowly three times, is first 
covered with glacial acetic acid for 5 seconds. The excess of the acid 
is then soaked up with filter paper and the rest washed off with aniline- 



THE SPUTUM: PLANT PARASITES 33 

water-gentian- violet (see page 281) renewing the stain repeatedly until 
all the acetic acid is removed. This stain is left on for about 3 min- 
utes. The film is now washed in an 0.85 to 2% aqueous solution of sodium 
chloride until the stain is washed off. The specimen is examined at once in 
this fluid. 

Very fair capsule stains may also be obtained by simply staining the 
smear for about 30 seconds with the ordinary aqueous gentian- violet 
solution (5 c.c. sat. ale. sol. to 95 c.c. of distilled water), then washing 
with a 1% potassium carbonate solution and studying the specimen 
in this fluid. 

Buerger's Method. 29 — The spread, before it is completely dry, is covered with 
Zenker's fluid minus acetic acid (bichromate of potassium, 2.5 gms.; sodium sulphate, 
1.0 gm.; water, ioo c.c; bichloride of mercury till the fluid is saturated, i.e., about 5%) 
and gently warmed over a small flame for from 3 to 5 seconds. It is then washed rapidly 
in water, flushed once or twice with alcohol (95%) and covered for from 30 to 60 seconds 
or longer with tincture of iodine (about 7%). The specimen is next washed with alcohol 
(to remove the iodine) until the alcohol remains clear and then the specimen is dried 
in the air. It is then stained for from 3 to 5 seconds with freshly prepared aniline-oil- 
gentian- violet (aniline oil 10; water 100; this mixture is shaken and filtered and to the 
filtrate are added 5 c.c. of saturated alcohol solution of gentian violet). The excess of 
stain is removed with a 2% NaCl solution. The spread is examined in this fluid. 

For methods of* cultivating Micrococcus pneumonia the reader is 
referred to Buerger's article. 30 In clinical work the mass of the sputum to 
be examined may be washed in several changes of sterile salt solution to 
free it of adherent mucus and then smeared on the surface of slants of 
Loffler's blood-serum. From this tube at the time of inoculation other 
tubes should be inoculated in sequence to secure some isolated colonies. It 
is seldom necessary to make more than 3 or 4 such dilutions. The rabbit 
and mouse are very susceptible to this organism, the latter especially. 
Its virulence is tested by inoculating a mouse with fresh sputum. With 
virulent organisms and a susceptible animal the pneumococcus can be 
recovered from the heart's blood and the spleen. This is by far the 
quickest and surest method of identifying this organism. A little of the 
sputum (or culture, or pleural fluid, etc.) is injected either subcutaneously 
or within the peritoneum of a mouse or rabbit. If this organism is present 
the animal will die in from 24 to 48 hours of septicemia and in smears of 
the beast's blood these encapsulated diplococci can be found in abundance. 
Considerable variation in virulence will be noted. 

Cole has shown that the organism formerly called Micrococcus pneu- 
moniae (Diplococcus lanceolatus, Diplococcus pneumoniae) is in reality a 
group of organisms consisting of at least 4 types which can be separated 

29 Report of the Medical Commission for the Investigation of Acute Respiratory 
Diseases of the Dept. of Health of the City of New York; Part I, Studies on the Pneu- 
mococcus, 1905. 

30 The Journal of Experimental Medicine, 1905, vol. vii, No. 5. 

3 



34 



CLINICAL DIAGNOSIS 



by biological tests. Their relative frequency and virulence are best illus- 
trated in the following table : 31 



Percentage 
incidence 



Mortality of 
cases not specifi- 
cally treated 



Found in the 

mouths of healthy 

persons 



Pneumococcus, Type I 

Pneumococcus, Type II . 

Pneumococcus, Type III 

(Pneumococcus mucosus) 
Pneumococcus, Type IV 



33% 

33-5% 

13% 

20% 



25% 

29% 



12.5% 
Sometimes 6% 



0.8% 

:8.2% 



52.9% 



The organisms of Type I are alike in so far as their immunity reactions 
are concerned. For this type an antiserum has been obtained, the use of 
which has reduced the mortality of cases thus treated from 25 to 8%. 
Type II also is a fixed type but the serum for the treatment of the cases 
with this infection is not nearly as successful. Pneumococcus mucosus 
(Type III) is a far more virulent organism against which no serum has 
as yet been obtained. This organism has larger capsules than do those of 
these other groups and forms a sticky exudate in animals. These three 
types are fixed types. Type IV is composed of organisms which have no 
common immunity reactions and which are much less virulent. The pneu- 
mococci found in the mouths of from 80 to 90% of normal persons are 
usually of this type. 

For the rapid isolation of the organism from the sputum for agglutina- 
tion tests to determine the type, a mass of the sputum is washed several 
times in sterile salt solution, rubbed up in a mortar with one-half a cubic 
centimeter of bouillon and injected into the peritoneal cavity of a mouse. 
Six hours later some of the peritoneal fluid is removed, or after about 
8 hours the animal is killed and the peritoneal exudate washed out with 
5 to 6 c.c. of salt solution or bouillon. This suspension of bacteria and 
leucocytes is then centrifuged at a rate sufficient to throw down the leuco- 
cytes. The supernatant fluid is withdrawn and centrifuged rapidly to 
collect the bacteria. The dense emulsion of bacteria thus obtained may 
be used for the agglutination tests. These are made by determining 
whether or not Serum I or II will agglutinate the organisms found. 
Often the type may in this way be determined at once. 

Friedlander's Bacillus. — This organism, known also as Bacillus 
mucosus capstdatus appears in the sputum and blood as a short, rather 
thick, encapsulated, non-motile bacillus with rounded ends. It occurs 
single, in pairs or in short chains. It does not produce spores. It is de- 
colorized by Gram's method. It grows on ordinary media. Injected into 
laboratory animals it produces a fatal septicemia. 

Bacillus influenza (see Fig. 14) is one of the smallest of the micro- 
scopically visible bacteria. It is a short, slender, non-motile bacillus, 

31 Trans, of the Assoc, of American Physicians, 19 15, vol. xxx, p. 234. 



THE SPUTUM: PLANT PARASITES 35 

with rounded ends, and has a tendency to grow into filamentous forms. 
It stains faintly and usually has a marked tendency to polar staining, so 
that it often resembles a diplococcus. It is decolorized by Gram's method 
and grows only on media containing hemoglobin. The most profuse 
growth is obtained if pigeon's blood is used. Its growth and virulence are 
increased if it is grown with other organisms. In the sputum it occurs 
free or in groups which are often large, while others are found in leucocytes. 
(Some think that increased phagocytosis is a sign of improvement.) Bacil- 
lus influenzae is sometimes virulent to animals (specially the guinea-pig 
and small rabbits) but often is not. 

The ability to recognize this organism in the sputum depends chiefly 
on one's familiarity with its morphology. A good method of staining the 
specimen is as follows : the fixed smear is stained with aniline-oil-gentian- 
violet (Sterling's) for i% minutes and washed in water; covered with 
Gram's solution i% minutes and again 
washed in water; immersed in 95% 
alcohol 5 minutes and washed in water ; 
covered with 0.2% aqueous Bismarck 
brown (20 c.c. of saturated alcoholic 
solution of Bismarck brown diluted 
with 80 c.c. of water) 1 minute, washed, 
dried and mounted. 

Cultivation of Bacillus Influenza — 
Avery s Oleate-hemoglobin Medium. 32 — 

Preparation Of the medium. Meat Fig. 14.— Sputum of influenza stained with 

. _ , _. . 1 . , . 1 . Gram's and Bismarck brown, showing the 

lniUSlOn agar (2%, See belOW) Which IS Bacillus influenzae (brown), Micrococcus pneu- 

, ....... .. .. • ,. monias, et al. (blue). X QOO. 

neutral or slightly alkaline in reaction 

is used as a base. To this is added a solution of sodium oleate sufficient to 
make a final concentration of 1 : 1000. A serum-free suspension of red 
blood- cells in broth is freshly prepared from sterile defibrinated rabbit's 
blood. One cubic centimeter of this corpuscular suspension is added to 
each 100 c.c. of oleate agar, the addition of blood being made while the 
medium is still hot. Plates are then poured, each containing about 15 c.c. 
of the oleate-hemoglobin agar and are used fresh to avoid the drying out of 
the medium. In the preparation of oleate-hemoglobin agar attention should 
be given to certain details: 

1. Agar. — Two per cent, meat infusion agar having a reaction of from 0.3 to 0.5 
acid to phenolphthalein should be used. The initial hydrogen ion concentration of the 
agar should represent a pH of from 7.3 to 7.5. Hormone agar prepared according to the 
formula of Huntoon 33 yields excellent results. 

2. Sodium Oleaie. — Two per cent, solution of sodium oleate (neutral) is made in 
distilled water, sterilized in the autoclave and kept as a stock solution. Five c.c. of 
this 2% solution of oleate is added to 95 c.c. of agar, giving a concentration of I : iooo. 

32 Avery, Jour. A. M. A., 191S, vol. 71, p. 2050. 

33 Jour. Infect. Dis., 1918, 23, 169. 



- i - ' - .. 






**>|W™ 




mm %.""> 


••^'^>f 


, , V ** 




• ' • N ;^.- n 






"SSL 


}^-" \ *\] ' * 


- ^ 




••*"" 


£<><&{**■ OOO 



36 CLINICAL DIAGNOSIS 

In the present work Kahlbaum's sodium oleate has been used, but other preparations 
are serviceable. 

3. Suspension of Red Blood-corpuscles. — Sterile defibrinated rabbit's or human 
blood may be used. Since serum is known to inhibit the action of oleate and since 
hemoglobin is the constituent of blood essential for growth of Bacillus influenzae, a 
serum-free suspension of the red corpuscles is used instead of whole blood. The red cells 
are removed from the defibrinated blood by centrifugalization, the supernatant serum is 
pipeted off and the corpuscles are made up to the original volume of blood by the addi- 
tion of broth. One cubic centimeter of this suspension of red cells is added to each 
100 c.c. of oleate agar. The suspension of blood-corpuscles should^ be added directly 
while the medium is hot and just before the medium is to be used. 

4. Formula. — This calls for: 

Agar 94 c.c. 

2% solution of sodium oleate 5 c.c. 

Suspension of red blood-cells . 1 c.c. 

Oleate-hemoglobin bouillon may be prepared in the same way by sub- 
stituting broth in the place of agar in the foregoing formula. 

Cultures taken from the nasopharynx by the West tube or from the 
throat by direct swabbing should be streaked on the surface of the oleate- 
hemoglobin medium according to the technic described for the detection 
of meningococcus carriers. 34 Similar plates should also be made from the 
sputum. At necropsy, cultures should be taken from scrapings of the 
tracheal and bronchial mucosa, as well as from the lung exudate. 

All culture plates should be incubated for 48 hours at 37 C. 

Pfeiffer's bacillus is an ubiquitous organism. To explain this its friends 
maintain that it has persisted with diminished virulence in areas formerly 
visited by epidemic influenza. Lord found it in 60 of 100 cases of non- 
tuberculous pulmonary infections with cough and in 29 of these in pure 
culture. This has been confirmed by many others. 35 It has been isolated 
from a surprisingly large number of cases of chronic bronchitis, asthma, 
tuberculosis, etc. The duration of these cases had varied from months 
to years, and one probably dated back for forty-five years. 

It occurs in bronchopneumonia as a primary and as a secondary {e.g., 
in diphtheria) invader. It is reported as a very common secondary invader 
in pertussis, after the paroxysmal stage begins (Bordet's bacillus?), in the 
bronchitis of measles, in lobar pneumonia and in tuberculosis with cavity 
formation. But what is more important it has been reported as the pri- 
mary cause of acute bronchiectasis, of cholecystitis, arthritis, pyelitis, 
cystitis, otitis media, acute nasal infections, empyema, meningitis, endo- 
carditis and of general septicemia. 

Not only is Bacillus influenzae the cause of conditions which in no way 
resemble epidemic influenza, but even the most typical of grippe epidemics 
have been shown to be due to quite other organisms than this; to Micro- 

34 Standard Technic of Meningococcus Carrier Detection, Adopted by the Medical 
Departments of the United States Army and Navy, 19 18. 

35 Holt, Arch. Int. Med., 1910, v, 449; Leutscher, Arch. Int. Med., 1915, xvi, 657; 
et al. For very complete bibliography see Arnold, The Jour, of Lab. and Clin. Med., 
July, 1920, v, 652. 



THE SPUTUM: PLANT PARASITES 37 

coccus pneumonias, Streptococcus pyogenes (hemolyticus and viridans), 
Streptococcus mucosus, etc. 36 Of a group of 31 cases of measles which 
developed a few weeks after an epidemic of influenza this bacillus was 
found in the sputum of 25. 

The opportunities afforded in 1918-20 to study the relation of this 
organism to the pandemic influenza were improved by many of the best 
bacteriologists of the world and their results have on the whole demoted 
this organism to a subordinate position. It certainly does not cause the 
primary infection called influenza or " flu " ; it is one of the earlier secondary 
invaders, an important member of the second of the 3 or 4 "crops" which 
follow each other ; while it may cause some but not all of its complications 
it would seem to have a definite place in the series of infections preparing 
the way for others, the various streptococci and pneumococci. So far as 
the influenza pneumonia is concerned the reports from different camps 
varied much, showing marked regional differences in the disease. 37 During 
an epidemic in Nashville, Tenn., 1920, Arnold 38 found Bacillus influenzas 
in 3 5 % of normal throats, in 7 7 . 7 % of the cases of acute rhinitis and pharyn- 
gitis and in 86.5% of the cases of influenza. 

Bordet's bacillus, the generally accepted cause of pertussis, is found 
early in pure culture in the tough masses of mucus and especially in the 
shreds of mucus from the smallest bronchi. It is decolorized by Gram's 
method and shows marked polar staining. Morphologically it resembles 
Bacillus influenzas, although as seen in the sputum it is rather longer and 
plumper than is this "but as a result of cultivation it becomes smaller 
and smaller until it frequently appears as a mere point under the highest 
powers of magnification" (Bordet). This organism does not grow on ordi- 
nary media, but on media which are weakly acid and poor in nutrient 
constituents and which contain ascitic fluid or blood. The first growth 
is seen in two or three days. It can be trained to grow on media which do 
not contain blood. Bordet advises the following medium: 100 gms. of 
potato are cut into small slices and mixed with 200 c.c. of water contain- 
ing 4% of glycerin and then heated in an autoclave. The fluid is then de- 
canted. To 50 c.c. of this extract of potato are added 150 c.c. of a 0.6% 
solution of sodium chloride and 5 gms. of agar-agar. This is then auto- 
claved. While warm it is filtered into test-tubes, 2 or 3 cm. into each 
tube, and sterilized. Blood (preferably human, although guinea-pig's blood 
will do), is defibrinated and added to the agar of each tube in equal 
quantities. The tubes are then shaken and slanted. 

Bacillus diphtheria or the Klebs-Loffler bacillus (Fig. 15), is a 
small straight or slightly curved rod, from 1 to 6, average 2 to 3, microns 
in length. It is a non-motile, non-liquefying, non-spore-producing aerobe. 

36 Davis, Arch, of Int. Med., vol. xi, No. 2, p. 124. 

37 See Opie, et al. f Jour. A. M. A., 1919, lxxii, 108. 

38 Loc. cit. 



38 CLINICAL DIAGNOSIS 

It grows on all ordinary culture media, providing they are not acid nor 
too alkaline, but best on blood-serum. 

One of the best media for this organism is Lbfflefs blood-serum. This 
is a mixture of blood-serum, three parts, and bouillon containing i% of 
glucose, 2 parts. It is coagulated at about 70 C. When no tubes of media 
are at hand, cultures can be made on the coagulated white Of a hard-boiled 
egg. The shell is lifted at one end of the egg, the surface inoculated and 
the shell put back. The egg is then put in a thermostat. 

Bacillus diphtherias grows with great rapidity on the above-mentioned 
serum. At the end of even 8 hours in some cases the growth can be seen 
and the organisms easily found in smears, but a negative examination at 
that time has no value. In any case the growth can be determined in 
18 or 20 hours. Until this time the diphtheria bacillus if present will 
have dominated and smears from the surface of the serum will look like 
m ssea ~*^x, ■■■*-— -jp^f^ pure cultures. From this time on, 
f^ I? £rf *tf >£*& *I however, the ordinary organisms from 

r *S J$ the throat begin to dominate and will 

_^X*< soon crowd out the diphtheria bacil- 

k . * lus, unless the latter happened to be 

present in pure culture. The colonies 

of Bacillus diphtherias on blood-serum 

are moderate in size, elevated, of a 

j •••.'. 1 grayish-white color and with opaque 

£— ■•--< ■■....' ' — d centers. 

Fig. 15.— Bacillus diphtheria;, from a young TV.io Ar „ otl ; cni If | Q 1 /otl /li-r^/M-'hr 

blood-serum culture. Photomicrograph by J- ™S Organism, it taxen OireCtlV 

Dr. Thomas M. Wright. from ^ throat Qr from & yQung 

culture (less than 24 hours old), presents when properly stained an almost 
characteristic appearance. Smear preparations for microscopic examina- 
tion are made by scraping some of the growth from the surface of the 
serum with a platinum needle. This is then rubbed on a clear glass slide, 
allowed to dry in the air and the slide passed through the flame 3 times 
in order to fix the specimen. 

A good stain for this organism is Loffler's Methylene Blue (saturated 
alcoholic solution of methylene blue 30 c.c, and aqueous solution of 
1 : 10,000 KOH 100 c.c). The slide, covered with this stain, is warmed 
for a few minutes (it will not overstain), the stain then washed off in run- 
ning water and the smear dried with blotting paper. The specimen can 
be improved by washing it further with 0.1% acetic acid, but this is 
seldom necessary. 

Neisser's stain, intended to differentiate this organism from all others, 
has been adopted by many as the standard stain. The specimen is stained 
for about 5 minutes with a methylene blue solution (methylene blue, 
Grubler, 1 gm., 96% alcohol 20 c.c, distilled water 950 c.c, glacial acetic 
acid 50 c.c; this stain is filtered before using). During the staining the 



THE SPUTUM: PLANT PARASITES 39 

dye should be frequently renewed and the specimen gently heated. The 
stain is then washed off with water. It is next stained in Bismarck brown 
solution for 2 minutes (Bismarck brown 2 gms. dissolved in 1000 c.c. of 
distilled water). The polar granules are stained a deep blue and the 
bodies of the bacilli a light brown color. 

Bacillus diphtherias is not decolorized by Gram's method, that is, it 
is a "Gram positive" organism. 

Gram's Stain. — The smear is stained for 1% minutes with aniline- 
gentian- violet (saturated alcoholic solution of gentian violet 5 c.c, aniline 
water 100 c.c; the aniline water is made by slowly mixing aniline oil, 
1 part, with distilled water, 20 parts; the mixture is allowed to stand 
for some hours and then filtered until clear). It is next washed in water 
and then put for 1 or 2 minutes in Gram's or Lugol's iodine solution 
(iodine 1 part, potassium iodide 2 parts, water 300 parts). The specimen 
is then washed in absolute alcohol for from 3 to 5 minutes. It may now 
be mounted, or it may be counterstained with Bismarck brown, washed, 
dried, and mounted. Bacilli which "stain by Gram," or are "Gram 
positive" retain the gentian- violet color. Those which "decolorize by 
Gram" or are "Gram negative" will be unstained unless a counter-stain 
is used. 

Diphtheria bacilli stained by the above methods show at their ends or 
along their bodies deeply staining blue granules called "polar granules." 
Some bacilli contain so many such granules that they have a beaded appear- 
ance. This irregularity in staining as shown by the polar granules or the 
beaded appearance depends in part on the age of the growth and may 
entirely fail. These bacilli vary also much in size. Some recognize a 
" long " form and a "short " form and think these forms differ in virulence. 
But no relation between length and virulence has been determined. The 
length of the bacilli seems to depend rather on the stage of the disease 
when the culture was made and on the age of the growth. 

For routine laboratory examinations, smears are made and blood- 
serum tubes are inoculated at the same time from material obtained directly 
from uhe throat or from the swab used. Smears are made from the culture 
before the growth is 24 hours old. If typical appearing bacilli are found 
in either of these two sets of smears, both stained by Neisser's method, 
a positive diagnosis is made. The smears made directly from the throat 
may show diphtheria bacilli and no growth be obtained, and frequently 
the growth will succeed when the search over the first smears was negative. 

One searches the smears for bacilli about 5/x long with brown bodies 
and 2 blue staining polar granules, 1 at each end. If an}^ answering this 
description are found a positive diagnosis of diphtheria is made. Hosts 
of other shapes and forms may be seen, but the presence of this form is 
considered conclusive. The barred and beaded forms are not as common 
as these with the two polar granules which are present in over 90% of 



40 CLINICAL DIAGNOSIS 

the cases. If the culture is over 24 hours old when examined these forms 
may not be seen and such a culture should be discarded. One may then 
find a host of involution forms with their bizarre shapes: spindle, pear, 
dumb-bell, lancet, club, and the varicosed forms. These may be found 
in smears made directly from the throat. They cannot be differentiated 
from the involution forms of other mouth organisms. 

If the organism were to be cultivated for several generations, and for 
diagnostic purposes this is never done, it would rapidly lose its character- 
istic morphology and could not be accurately differentiated by its morphol- 
ogy alone from other bacilli. For these reasons the routine described 
above is accurately followed in making clinical laboratory examinations 
for diphtheria and if typical forms are not found a fresh bacteriological 
examination is made. The statement is often made that Bacillus diph- 
therias can be found in the throats of healthy persons who have not been 
exposed to diphtheria. McCollum states that Lofner found it in 4 of 160 
such individuals, Park and Beebe in 8 of 330, Kober in 5 of 600 and Denny 
in 1 of 235, but that he could not find it in any of the 130 such persons he 
examined. McCollum doubts also that it is often found in the throats of 
healthy persons who have merely been exposed to this disease, since he 
failed to find it in the throat of a single one of 60 nurses from the diphtheria 
wards of the Boston City Hospital. Some of the earlier reports of the 
presence of diphtheria bacilli in the throats of normal individuals may 
have been made before the question of pseudo-diphtheria bacilli had 
arisen, but it is also possible that some of these so-called normal persons 
had had a mild diphtheria, for some diphtheria patients do not realize 
that they are ill, that is, that they have a definite inflammation due to 
this bacillus in their throat or nose. It is also true that Bacillus diphtherias 
can live for a long time in the throats and noses of those who have recovered 
from a latent attack of diphtheria. 

The test of virulence is sometimes important for the recognition of the 
diphtheria bacillus. To make this test a guinea-pig is inoculated subcu- 
taneously with a bouillon culture not over 48 hours old of the organism 
to be tested. The virulence of Bacillus diphtherias tends to diminish after 
2 days of growth. The amount of the bouillon culture medium injected 
should equal 1% of the animal's weight. If the organism injected is 
Bacillus diphtherias, the animal will soon show symptoms of acute or of 
chronic infection. If acute, the animal will die in from 1 to 6 days, and at 
the seat of inoculation will be found extensive necrosis with a marked in- 
flammatory reaction. There will be extensive edema of the abdominal 
wall, effusions into the serous cavities, hemorrhages into the adrenals, 
swelling of and hemorrhages into the lymph-glands and focal necrosis in 
various other organs. 

If a chronic infection results, the animal will show paralysis similar 
to that in man, and will die in about 6 weeks. 



/ fe » 



THE SPUTUM: PLANT PARASITES 41 

Formerly all organisms with the morphology described above were 
considered Bacillus diphtheria. Since then various pseudo-diphtheria 
bacilli have been described, and the relation of these to Bacillus diphtherias 
is still a mooted question in bacteriology. 

The following groups of organisms may be mentioned : 
i. A bacillus whose morphology is typical, with typical cultural char- 
acteristics, especially the ability to form acid from glucose, and which 
produces the typical lesions in animals, is, in the opinion of all observers, 
Bacillus diphtherias. 

2. Bacilli with typical morphology and typical cultural reactions, espe- 
cially the ability to form acid from glucose, but which are not pathogenic 
to animals, may be called avirulent diphtheria bacilli. Roux maintains, 
however, that the ability of the organism to ferment the sugars is not an 
essential characteristic of the species. 

3 . Bacilli whose morphology is typi- 
cal but which do not conform in their 
cultural reaction with the diphtheria 
bacillus and which are either non- 
pathogenic to animals, or do not pro- 
duce typical lesions, may properly be 
called pseudo-diphtheria bacilli. 

4. Finally, there are a number of 
organisms which resemble Bacillus 
diphtherias in many ways, but whose 
morphology is not the same. 

r °- / Photomicrograph by Dr. H. Schapiro. 

Bacillus Fusiformis and Vincent's 
Spirocheta (Fig. 16). — Bacillus fusiformis (Vincent) is a long slender bacil- 
lus often fusiform in shape, indeed the ends may be quite pointed. This 
bacillus is straight as a rule, but many are curved and a few may be S- 
shaped. It is non-motile (disputed), it decolorizes by Gram (disputed), is 
often beaded, and can be grown in pure culture. 39 It is best stained 
by carbolfuchsin. 

Carbolfuchsin. — This stain is made up of basic fuchsin 1 part, absolute 
alcohol 10 parts, 5% carbolic acid 100 parts. The specimen may be covered 
with this concentrated stain for about half a minute, or, better, this stain 
is diluted with from 5 to 10 times its volume of water and then left on the 
smear for about 5 minutes. 

This organism has been found in many normal mouths, on the surface 
of normal tonsils, in the cavities of decayed teeth, in the exudate of pyor- 
rhea alveolaris, in antrum disease, aphthous ulcers, and also in fetid 
abscesses elsewhere in the body. This organism is common enough 
but has escaped notice since it is cultivated with difficulty and when 
seen in smears is usually passed by as a "harmless saprophyte." Recent 

39 Weaver and Tunnicliff, Jour, of Infect. Dis., 1905, ii, p. 446. 







\ v ^* , ^ s - "^vjB 


X 
i 




/ / "* *£• 


1 






1 




" N *& 


1 




; 




W 


\ 1 u, 
1 • J 


Fig. 


16. 


— Smear from the throat of Dr. Louis 


P. 


Hamburger's case of Vincent's angina. 



42 CLINICAL DIAGNOSIS 

work, however, is rather in favor of the view that this organism is patho- 
genic and pyogenic. 

In the mouth this organism is often, but not always, associated with 
a spirillum or spirocheta, also called the Spirocheta of Vincent, which 
measures from 15 to 25/z in length, is twisted in from 2 to 5 spiral turns, 
is actively motile, is Gram negative and stains so faintly that it is often 
overlooked. It has not yet been cultivated. This is quite certainly a 
saprophyte which occurs in enormous numbers in the mouths of even 
healthy persons and yet it is so frequently associated with Bacillus fusi- 
formis that the two are supposed to be symbiotic and together cause 
the ulcers of Vincent's angina. 

Sinclair (see page 59) believes that these organisms are important 
in explaining some of the hemorrhages of incipient tuberculosis. He 
examined the sputum for these organisms fresh and also cultured in 
broth under albolene and claims to get in 72 to 96 hours a growth of these 
dual forms. 

Micrococcus tetragenus is a micrococcus which occurs always in tetrads, 
the individuals a little larger than the ordinary staphylococcus, their adja- 
cent surfaces flattened and the group surrounded by a thick mucous capsule. 
Occurring usually in the mouth as a saprophyte it is, however, sometimes 
pathogenic and is found in the sputum in cases of bronchitis, tuberculosis 
with cavity formation and in hemorrhagic infarction. It is supposed to 
aid Bacillus tuberculosis in its destructive processes. Some would separate 
a pathogenic variety, which can be cultivated, from the common, harmless 
mouth saprophyte which will not grow on laboratory media. 

SarcincB are seldom found in the sputum. One does occasionally find 
them in cases of gangrene, tuberculosis, bronchitis (see page 72), pneu- 
monia, and in the sputum of old, debilitated persons in whom they often 
are the cause of the gray patches of stomato-pharyngomycosis sarcinia. 
They probably are harmless saprophytes. 

Pathogenic yeasts are much more important causes of pulmonary 
lesions than is generally believed and may explain many of the cases of 
chronic lung trouble under treatment for tuberculosis. Busse was the first 
to call attention to these organisms. In his case of " saccharomycosis 
hominis" due to Saccharomyces busse the original infection was of the 
tibia, but later there developed in both lungs caseous cavities in which 
the yeast was present. The yeast cells were rather small, averaging about 
SfjL in diameter, oval, very refractive and resembled fat droplets except for 
their greenish shimmer. The younger cells were homogeneous, but the pro- 
toplasm of the older ones is granular and the nucleus visible. They are 
made clearer by the addition of sodium hydroxide. Dr. Breed 40 states 
that after her attention had been directed to the presence of these organ- 
isms in the sputum of cases supposedly tuberculous she made cultures of 

40 The Arch, of Int. Med., August 15, 1912, vol. x, No. 2, p. 108. 



THE SPUTUM: PLANT PARASITES 43 

all sputa in which on careful examination no tubercle bacilli had been 
demonstrated and in less than 2 years found at least 10 cases of possible 
pulmonary saccharomycosis. Of course yeasts are not rare as secondary 
invaders in tuberculosis. To find them the sputum is washed in tenth- 
normal sodium or potassium hydrate and cultures made from the particles 
resembling pus. Dr. Breed grew these yeasts on practically all the common 
media and tested the pathogenicity of the organism to rabbits, white mice, 
guinea-pigs and monkeys. 

A patient was admitted to the Indiana University Hospital with many sinuses of 
several months' duration on the upper anterior chest wall, pus discharging. Some of 
these certainly led to the deeper tissues including the costal cartilages. Almost pure 
cultures of a slowly growing yeast were obtained from the sputum as well as from pus 
aspirated from the deeper portions of the sinuses. 

Blastomycosis. 41 — Blastomycetes (Fig. 17) are budding protophytes 
belonging to the same group as yeasts (Fig. 71, page 363). These organ- 
isms, round or oval in shape and from 8 to 12/z in diameter, have a finely 
granular often vacuolated protoplasm and a capsule with double contour 
separated from the protoplasm by a clear zone, often wider on one side 
than on the other. Reproduction in the living tissue is by budding only; 
in cultures, by mycelium formation. These organisms are pathogenic to 
rabbits, producing general systemic infection if injected intraperitoneally 
or intravenously. Subcutaneous inoculations are usually unsuccessful. 
They grow well on glycerin, glucose agar, blood-serum, bouillon, and other 
ordinary media. The growth at room temperature is microscopically visible 
in from 2 to 14 days, is dry, and develops many aerial hyphas; that in the 
incubator is pasty and moist, with fewer aerial branches. 

Thus far a few cases of general blastomycosis in man have been reported, 
the majority among foreign-born patients in Chicago. The symptoms 
suggest tuberculosis and the correct diagnosis has seldom been made 
before the subcutaneous abscesses had appeared. In practically every 
case the lung is involved sooner or later; indeed, evidence would indicate 
that in at least 65% of all cases the primary infection is in the lung. The 
organism may be demonstrated in the blood, urine and sputum. In the 
sputum one finds these cells in enormous numbers. It is probable that 
they are present in the sputum very early and yet unfortunately the diag- 
nosis has not yet been made from sputum examination. The general 
character of the sputum will depend on the pulmonary lesion, whether 
there is present bronchitis, bronchopneumonia or frank pneumonia, small 
metastatic abscesses, cavity formation or fibroid changes with or without 
dilatation of the bronchi. While the characteristic sputum is abundant 
and very blood-stained, the patients may for long periods expectorate 
only clear mucus or a mucopurulent sputum. 

41 See Montgomery and Ormsby, Arch. Int. Med., Aug., '08, vol. ii, No. 1, p. 1; 
also Fontaine, Haase and Mitchell, Arch. Int. Med., Aug., '09, vol. iv, No. 2, p. ioi. 



44 



CLINICAL DIAGNOSIS 



C. K., No. 9391, aged 59, was admitted to the surgical ward March 25, 1920, for mul- 
tiple abscesses especially of the extremities, due to blastomycosis. He died April 10, 1920. 

His sputum was a clear, white, very ropy mucus which contained very many of 
these parasites (see Fig. 17) and no red blood-cells. At autopsy both lungs were found 
to be the seat of extensive bronchopneumonia which showed no evidence of secondary 
pyogenic infection. 

On March 27, 1920, the red blood-cell count was 4,000,000 and the hemoglobin 75%. 
The leucocyte count was 10,200, of which 8.3% were small mononuclears, 0.8% large 
mononuclears, 90.3% polymorphonuclear finely granulars and 0.5% eosinophiles. The 
morning urine had a specific gravity of 1.020 and 1.022, was straw-colored, turbid, 
contained a trace of albumin, no sugar, no casts, but very many yeast cells. 




Fig. 17. — Balantidium Coli photographed in Stool (after MacCarty; Barker's Clinical Diagnosis, 

D. Appleton & Co.) 



Coccidiosis — Coccidioidal Granuloma — California Disease. — 
Thus far but about 40 cases of infection with Oidium coccidioides or Coccidi- 
oides immitis have been reported, nearly all from the San Joaquin Valley, 
California. 42 This organism is not to be confused with blastomycetes and 
the clinical picture of this infection is different from blastomycosis. 

Oidium coccidioides, Coccidioides immitis, belongs to the same group 
as the yeasts and the blastomycetes. It differs from the latter in that, 
while the blastomycetes multiply in the living tissues by budding only, 
this multiplies solely by endosporulation. 



42 Hektoen, Jour. A. M. A., Sept. 28, 1907, vol. 49, p. 1071. 
are reviewed. 



In this paper 17 cases 



THE SPUTUM: PLANT PARASITES 45 

Most patients with this infection belong clinically to the pseudo- 
tuberculosis group. "In fact, this disease presents the best mimicry of 
tuberculosis ever seen" (Hektoen). In most cases the initial lesion would 
seem to be in the lungs, which at autopsy are studded with disseminated 
miliary tubercles, areas of bronchopneumonia, or pulmonary abscesses. 
The sputum is mucopurulent or blood-streaked and in several of the cases 
contained the organism. 

Oidium albicans is the very common parasite of thrush. This mem- 
brane can develop in the bronchi as well as in the mouth and pharynx. 
One sees it most often in the mouths of children, especially the weak 
babies, during their first week and of adults weakened by old age or 
disease, especially b}^ diabetes or typhoid fever. In these cases we may 
find this growth in the throat, nose, esophagus, bronchi and lungs. The 
most common form found is the large-spored variety which liquefies gelatin 
in culture rather than the variety which produces smaller spores and 
which does not liquefy gelatin. In the sputum may be found these cells 
which resemble ordinary yeast in all particulars. They are from 5 to 6ju 
long and 4//, wide, oval in shape. When subjected to unfavorable cultural 
conditions this organism may grow in threads of very variable size and 
length, with double contour, the protoplasm of which contains droplets, 
granules and vacuoles. In these threads develop true endogenous spores. 

Actinomycosis infection (Actinomycosis bovis) of man is rare in 
America. In about 15% of the cases reported the disease would seem to 
have been primary in the lung. Clinically, the picture suggests tubercu- 
losis, while the sputum presents no distinctive feature other than the 
actinomyces granules. Some cases resemble a miliary tuberculosis; some 
begin as a slight catarrhal bronchitis which soon develops into a subacute 
bronchitis, with mucopurulent sputum often blood-streaked. In most of 
the cases, however, a bronchopneumonia is present from the first. These 
consolidated areas break, down forming cavities which contain fluid, pus, 
fatty detritus, fat globules, degenerated red blood-cells and the sulphur 
granules. The ulcerating process progresses steadily through the lung 
and pleura to the chest wall which it, perforates. This disease is slow in 
its course and progresses without periods of improvement until death. 
The sputum may at first be scanty and odorless, consisting of pure mucus, 
but in most cases it is mucopurulent or purulent and often (in about 
one-half the cases) hemorrhagic and fetid. It is sometimes as rusty as 
in pneumonia, sometimes resembles current jelly, while few cases have 
terminated in fatal hemorrhage. Some patients have expectorated at 
one time a large amount of offensive yellow material which in at least 
•one case gave the patient the sensation as if the mouth were full of sand 
which grated between the teeth from the presence of the granules (Lord). 
If the sputum be carefully watched a correct and early diagnosis can usually 
be made since the sulphur granules which one finds are characteristic of 



46 CLINICAL DIAGNOSIS 

this disease. They are small granules varying in size, some requiring the 
microscope for their demonstration, some even 2 mm. in diameter. They 
are round and have a yellowish, grayish, greenish, or brownish color. 
Sometimes one finds many in the sputum, sometimes few. Microscopically 
they consist of a network of fine, twisted threads radiating from a center 
and at the ends of which are the characteristic club-shaped swellings, 
which when present in large number form a ring around the granular 
central mass giving it a radiating or star-like appearance. 

It is a good rule in any atypical case of lung disease to exclude actino- 
mycosis. One must, however, not mistake masses of mouth leptothrix 
and masses of degenerating cells for this organism. 

Moulds are often found in sputum, since special precautions in collect- 

ing and handling the sputum 

would be necessary to avoid this 
common air and dust contamina- 
tion. Eliminating these accidental 
contaminations, one finds moulds 
*C^ • ,. in the sputum of those cases only 

who have destructive processes of 
the lung. Whether these "bron- 
chopneumomycosis " are primary 
or secondary has been a much 
disputed point. Virchow believed 
that mould infections were usually 
secondary but that they could be 
the primary infection in necrotic 
-^ tissue, for example in areas of 
hemorrhagic infarctions which 
they would transform into cavities. While one often finds a few moulds 
in tuberculous cavities, in others they are the prevailing organism.. The 
contents of such cavities are as a rule odorless. There would indeed 
seem to be such an antagonism between moulds and the bacteria of 
decomposition that a cavity filled with the former is protected against 
the latter and vice versa. It is possible therefore that what would seem 
to be primary mould infections are those in which the moulds merely 
aided other organisms in the necrosing process and then crowded out the 
primary invader. Recently, however, through the work of the French, 
also of Saxer and others, it seems clear that Aspergillus fumigatus can be 
the primary invader and cause by necrosis an odorless cavity. 
Among the pathogenic moulds are: 

(1) Mucor. — There are 130 varieties of mucor, 6 of chem definitely 
pathogenic. Mucor moulds are a very common air form. The group as 
a whole is characterized by the mycelial growth, which branches much 
and which at first is unicellular although later septa may develop, and by 






Fig. 18. — Mucor mucedo. X 60. 



THE SPUTUM: PLANT PARASITES 47 

its sporangia which develop at the end of erect hyphse and consist of a 
columella surrounded by masses of spores enclosed by a membrane. Fig. 18 
represents Mucor mucedo, a very common, harmless form. If a mucor 
mould be found in the sputum the observer should note carefully the shape 
of the columella, the size of the spores and the nature of the membrane, 
although for positive identification cultures are necessary. There are 
several varieties of mucor known to be pathogenic: Mucor corymbifer 
is a fine, delicate, small mould the spores of which are 2 by 3/j in size. 
Its sporangia are colorless and pear-shaped, varying in size from 10 to 70^, 
and its membrane transparent. The columella, evident only when the 
spores have dropped off, is colorless and shaped like a boy's top, the larger 
end distal. This mould is in man the most common cause of kerato-, oto-, 
pharyngo-, and pneumomy- 
cosis. The sporangia-bearing 
hyphas of Mucor rhizopodi- 
formis are single, or branch 
as in a sheaf, short and of 
a brownish color. The spo- 
rangia are globular, black 
when ripe and the membrane 
opaque and soluble in water. 
The columella is brownish, 
from 50 to 75m wide, is con- 
stricted at its base, which is 
also truncated and with a 
wide flat apophysis to the 

margin of which the mem- :"\ ; 

brane is attached. The spores „ . ... , . ' 

r Fig. 19. — Aspergillus fumigatus. X 300. 

are colorless, spherical and 

from 5 to 6jli in diameter. Mucor racemosus has spores from 5 to 8/x 
long and 4 to 5^ wide. Its columella is elliptical in shape. Mucor 
pulsillus has sporangia which are black, from 60 to 80/x wide and covered 
by a thorny membrane. The columellse are egg-shaped or spherical, light 
brown in color and from 50 to 6o/j, wide. The spores are very small, round, 
colorless and from 3 to 3 .5/x in diameter. Mucor septatus has a pale, grayish 
brown, spherical sporangium and small colorless columella which after the 
loss of the spores may grow still longer. The hyphse have septa, hence 
the name of this mould. Its spores are about 2.5,0, in diameter. Mucor 
racemosus has black sporangia which are 70/* in diameter. The membrane 
is transparent, the columella round, the spores colorless, opaque and from 
3 to 4fi wide and 5 to 6ju long. 

The above forms of mucor are known to be pathogenic; almost all of 
them have been demonstrated in the ear. It is interesting that in all 
literature only 4 cases are cited in which they have been demonstrated 



oca 



o 



48 CLINICAL DIAGNOSIS 

in the lung and, so far as we know, in none of these cases were they found 
in the sputum before death. 

Aspergillus Fumigatus (see Fig. 19). — This is by far the most important 
of the pathogenic moulds. Its mycelium is a thick mesh of threads from 
3 to 6/jl wide, the finest without, but the oldest with, septa. The conidia- 
bearing hyphce are short, club-shaped and from 8 to 10/x in diameter at 
the larger (distal) end. The sterigmata, unbranched and from 6 to 15/i 
long, radiate from a central point, thus giving the head a fan-like appear- 
ance. The conidia spores, a chain of which tips the end of each of the sterig- 
mata, are round, colorless, and from 2.5 to 3^ in diameter. The size of 
these spores is important since those of Aspergillus glaucus are from 7 
to 8/x in diameter. All parts of this mould have a color which varies from 

_____ brown to dark grayish green. 

The spores can be found almost 
anywhere in nature, as one 
may demonstrate by exposing 
a moist piece of bread to the 
air for only a few minutes and 
' ' ^.^o °§£ then placing it in the thermo- 
stat. Aspergillus flavus (see 
Fig. 20) has conidia-bearmg 
^ r .jc o hyphas which are from 7 to iOyu 
thick. The young head has a 
yellowish or green color ac- 
cording to whether it is dry or 
wet and is brown when old. 
£JM The conidia themselves are 
round, of a sulphur yellow 
color and from 5 to 7^ in diameter. Aspergillus niger is of a chocolate 
brown color and has conidia from 3.5 to 5/x in diameter. The growth 
of Aspergillus subfuscus is of an olive-green to a black color. It 
strongly resembles Aspergillus fumigatus, but is more pathogenic. Of 
all moulds Aspergillus fumigatus is the only one that has been shown 
to cause a primary lung infection. Sticker has collected from literature 
20 cases in which no other disease of the lung was present. In 16 of 
these Aspergillus fumigatus was found, in 4 cases the mould was doubt- 
ful. One of these 4 cases, reported by Osier, was a woman who for 12 
years had expectorated masses of mycelium the size of a bean, grayish, 
and of a downy consistency. An interesting case of primary chronic 
"membranous" bronchitis due to Aspergillus fumigatus was reported 
by Devilliers and Renon. 43 The patient was a grain sorter. Frag- 
ments of membrane composed of the mycelium of this mould (recog- 
nized from cultures) were expectorated monthly. These casts were from 
43 Le Presse Med., 1899, ii, p. 325. 



".-; 



Fig. 20. — Aspergillus flavus. X 300. 



THE SPUTUM: PLANT PARASITES 49 

i to 6 cm. long and, having no branches, probably originated in the 
larger bronchi. 

Pneumomycosis Aspergillina. — Sticker u has divided the cases of asper- 
gillosis into a "sporadic" group, which includes feeble patients susceptible 
because of their debilitated condition to such infection as well as persons 
suffering from other lung disease of which this is a secondary infection and 
an "endemic" group of cases who owe this disease to their occupation. 
Among the latter are the pigeon feeders, who are much exposed to the 
moulds of grain, and the hair combers, who work in an atmosphere so 
laden with infected dust that the cat is the only animal that can live 
with them. No autopsies on such cases have as yet been reported. 

Clinically, many of these cases resemble chronic pulmonary tuber- 
culosis and so form the pseudo-tuberculosis group of mould cases. The onset 
usually is with a hemorrhage, either slight or profuse, which recurs at inter- 
vals during the course of the disease. The cough at first is rather dry, then 
they raise a frothy sputum which often contains blood flecks. The sputum 
soon becomes greenish and purulent. Such sputum may be expectorated 
for months, even for years. Toward the end the expectoration is still green- 
ish but more purulent, nummular, and sometimes more hemorrhagic. 

In another group the process is a chronic bronchitis resulting in cirrhosis 
of the lung. The sputum of these patients is abundant, foamy, and watery. 
In Wheaton's case 45 the condition during life simulated actinomycosis, 
but at autopsy only a few tubercles were found and a large cavity. Some 
cases have expectorated casts of the bronchi made up of mycelium threads. 

W., No. 1031, aged 47, admitted May 2, 1915, is a good illustration of this chronic 
bronchitic form of pulmonary aspergillosis. That these cases may give a distinctive 
clinical picture is suggested by the fact that this patient, admitted for pulmonary 
tuberculosis, when first met at ward rounds was at once demonstrated to the students 
as a probable case of pulmonary aspergillosis since the subjective symptoms and the 
physical signs were out of proportion to the effect the disease had had on the patient's 
general condition. The man had been considered tuberculous for many years, had been 
admitted twice to a special hospital for the tuberculous, although every one of the 
numerous examinations of the sputum had been reported negative. His dyspnea in 
the recumbent position was so great that for 9 years he had slept in a chair and yet 
during much of this time he had been able to earn his own living and had always main- 
tained his weight. The widespread and marked physical signs of the lungs, the impaired 
resonance on percussion, the tubular modification of the breath sounds and the wide 
distribution of the rales which were moist and dry, coarse, medium and fine, suggested 
a very serious pulmonary condition and yet the patient had little or no fever and was 
able to live a fairly active life. The sputum contained the mycelial threads of Aspergillus, 
fumigatus. For several years this patient returned periodically for the benefit of our 
classes in physical and clinical diagnosis. 

While demonstrating him at ward rounds one day a visiting physician became much 
interested and said, "This must be what my wife has," which a later examination 
proved to be the case. (Mrs. K., aged 31, who had for years been considered a case 
Df tuberculosis.) 

44 Nothinagel's System, 1900, xiv. 

45 Trans. Path. Soc, London, vol. xliv, p. 38, 
4. 



50 



CLINICAL DIAGNOSIS 



For the diagnosis of pulmonary aspergillosis the mould itself must be 
demonstrated in the sputum. In fact, the sputum should be examined 
for moulds and yeasts in all cases of suspected tuberculosis in which 
Bacillus tuberculosis cannot be demonstrated. One may find in the fresh 
sputum the mycelium threads, the conidia hyphse, or the spores, yet the 
mycelial threads may not be found since they disintegrate rapidly. In 
dried and stained specimens one finds nothing suggesting a mould. It is 
important that the sputum of mould cases is usually odorless even though 
it contains large masses of lung tissue from a gangrenous lung. 

Penicillium glaucum (see Fig. 21), the most common of our media 
contaminations, has segmented condia-bearing hyphae which divide brush- 
like at the end, the branches being tipped .by sterigmata which are flask- 






Fig. 21. — Penicillium glaucum. X 300. 

shaped. The conidia are from 2 to 3 /z in diameter. This mould is non- 
pathogenic. Penicillium mummula certainly is pathogenic for animals 
and has been found in the ear of man. 

Although one can recognize in the fresh specimen the general class 
of a mould if one can find a sporangium, i.e., whether it is mucor, asper- 
gillus, or pencillium, yet to determine the sub variety cultures are neces- 
sary. This may be done by spreading the sputum over a piece of bread 
as media, or by using Sabourand's medium (maltose, 3.7; peptone, 0.75; 
and water, 100). 

The moulds may be stained in fresh specimen of sputum by a saturated 
watery solution of safranin or, better still, of thionin. 



ANIMAL PARASITES 



The infusoria found in the sputum are rarely important. Artault 
described Ameba pulmonalis as "a small ameboid cell which when stained 
looks exactly like a leucocyte, but while motile differs from it in its refrac- 



THE SPUTUM: ANIMAL PARASITES 51 

tility." Entameba histolytica (see page 401) is found in the sputum of 
cases of liver abscesses which have perforated through the lung. Enda- 
meba buccalis is found in abundance in the pus of cases of pyorrhea alveo- 
laris and so could of course be found in the sputum of such patients. This 
is probably the ameba found in the contents of abscesses of the jaw com- 
municating with the mouth. 

Flagellata have frequently been found in the sputum. A. Schmidt 
described such organisms in Dittrich's plugs; Artault, in the contents of 
a large tuberculous cavity ; others in the sputum cases of lung gangrene 
and putrid bronchitis. In one case of abscess of the lung following pneu- 
monia with operation 6 weeks after the onset of the pneumonia the sputum 
contained large numbers of flagellates. The name Trichomonas pulmonalis 



Fig. 22. — Sediment from echinococcus cyst. Above and to the left are two degenerated scolices 

( X about 60) ; to the right is the head of a scolex (X 400) ; below are hooklets of unusual shapes 

and a small mass of cholesterin crystals. X 400. 

has been suggested for these protozoa and yet, so far as we know, they are 
identical with Trichomonas vaginalis (see page 404). Cercomonads (see 
page 404) have been found in the sputum and in Dittrich's plugs. 

Echinococcus Disease. — Next to the liver the' lung is the organ most 
often (in 16.8%) infected with T&nia echinococcus (see Fig. 22). If a 
pulmonary cyst bursts, or one in a neighboring organ {e.g., the liver) 
ruptures through the lung, the sputum may contain daughter cysts, scolices, 
hooklets, or fragments of membrane from the cyst wall, any one of which 
is characteristic of the disease. 

The echinococcus cyst wall (see Fig. 23) consists of 2 layers, an external 
laminated articular capsule and an internal granular parenchymatous 
endocyst. The cyst content is a clear, limpid fluid which has a specific 
gravity of from 1.005 to 1.015. It is neutral or slightly acid in reaction 
and contains considerable sodium chloride, but no coaguable albumin. 
Inosite, leucin, tyrosin, succinic acid and hematoidin have been demon- 
strated in this fluid. From the endocyst, buds develop which become 



52 



CLINICAL DIAGNOSIS 



•3 



¥.C-:' 



smaller daughter-cysts and which break loose and lie free in the parent cyst. 
Inside these daughter cysts grand-daughter cysts may in turn develop. 
From the inner surface of the wall of any one of these cysts brood-capsules 
may form. These are cysts from the inner or outer wall of which the 
scolices grow. A scolex is the head of a Taenia echinococcus, and consists 
of a rostrum, which while alive it actively protrudes and retracts with 4 
suckers and surrounded by a circle of hooklets. Sooner or later the parasite 
dies and the cyst wall degenerates, its contents become an inspissated 
mass of cheesy material containing many free hooklets and dead scolices 
which later may receive a coating of calcium carbonate. 

These cysts in the lung often lead to pulmonary 
gangrene; the cyst may rupture and its contents 
become infected, or the infection of its contents may 
precede its rupture. In either case the result is a 
cavity or cavities connected with bronchi. 

The cough may at first be dry and hacking, and 
then as the cyst increases in size, some mucus may be 
expectorated. Hemoptysis is an early and important 
symptom. It may be the first indication of the dis- 
ease. Preceding rupture there are merely blood-streaks 
or flecks in the sputum, but even then hemorrhage may 
be profuse. Rupture of the cyst is often accompanied 
by a profuse hemorrhage which may be fatal. When 
a cyst ruptures its cavity soon becomes infected and 
the sputum then is fetid pus, sometimes of a chocolate 
color resembling the contents of an hepatic abscess. 
If the suppuration of the cyst contents precedes its 
rupture the cyst then becomes a closed abscess cavity 
full of pus and the symptoms of its rupture will be 
those of rupture of an abscess. As a rule, unless some 
of the characteristic cyst contents is found in the sputum, cases of this 
disease are diagnosed phthisis or pulmonary gangrene. If daughter cysts 
and pieces of membrane are found in the sputum it means that the cyst 
has opened into a large bronchus. In these cases the sputum may for 
months contain pieces of this membrane. 

Paragonimus Westermani. — This parasite, the "lung fluke," is the 
cause of the parasitical hemoptysis of man which is so common in Japan, 
parts of China, and Korea. In some mountain towns a majority of the 
people are said to be so infected. In Okayama 0.4% of all hospital cases 
admitted and in Kumamoto 5.9% of all pulmonary cases admitted showed 
this infection. 46 Stiles has reported 1 case in this country, that of a Japanese 
who had recently immigrated and who came under the care of Doctor 
Mackenzie of Portland, Oregon. A parasite which seems identical with 
46 Inouye, Zeits. f. klin. Med, 1903, vol. 1, p. 120. 




Fig. 23. — A small frag- 
ment of echinococcus 
cyst wall on cross frac- 
ture, showing transverse 
striation and pectination. 
X 50. 




THE SPUTUM: CHEMICAL EXAMINATION 53 

this had previously been found in domestic animals in this country and 
it is perhaps only a matter of time before more cases will be reported from 
among our patients with "tuberculosis." The duration of the disease is 
long, usually from 10 to 20 years from the appearance of the first symptom. 
The sputum is generally scanty in amount, very viscid, and consists of 
small pellets of blood mixed with mucus. In some cases it is as rusty as 
that of pneumonia. When no blood is present the sputum still may have 
any shade of yellow or brown, or dirty red due to the eggs themselves when 
very numerous. Colored spirals resembling somewhat Curchmann's spirals 
are quite characteristic. The diagnosis of this disease rests wholly with 
the discovery of the characteristic eggs in the fresh sputum. These may 
be expectorated in large numbers. They have a thick, smooth shell of a 
dirty reddish color with a characteristic lid at one end, which is not always 
evident, or not exactly on the end, and which may be partly shelled off. 
These eggs (Fig. 24) are from 66 to 96^ long and from 
48 to 6o/z wide. The amount of blood in the sputum is 
usually small, at the onset but a few drops, but later 
even from 300 to 800 c.c. may be expectorated in a few 
hours, especially if the patient leads a laborious life. 
In some cases these severe hemorrhages recur with 
great frequency, in others the disease takes a very slow 
course. The hemorrhage is always arterial. Its cause FlG .- 2 4-— Egg of Para- 

J gommus westermam 

is not clear because the ova are not contained in the from A/r th e sputum of 

Dr. Mackenzie s case 

pellets of the fresh blood. Charcot-Leyden crystals (t , hr ° u s h * he N kindness 

^ J J of Dr. Stiles). X 400. 

are common in this sputum, "sufficient proof that 

such crystals do not explain asthmatic paroxysms, since these cases 

never have asthma." 47 

Strongyloses Intestinalis. — Gage 48 reported a case of bronchopneumonia 
with larva of Strongyloides intestinalis in the sputum (see page 413). 

Ransom, 49 suggests that the larvae of Ascaris lumbricoides cause definite 
pulmonary symptoms when they pass through the lung from blood-stream 
to trachea and thence to the stomach and bowel. 

CHEMICAL EXAMINATION OF THE SPUTUM 

The chemical examination of the sputum offers but little of value in 
diagnosis and yet this little should not be disregarded. The determination 
oftenest made is that of the relative amounts of mucin and albumin. The 
test to be recommended is Zeonts modification of Schmidt's method, a micro- 
chemical test. . A small mass of the sputum is spread on a cover-glass, 
treated for at least a quarter of an hour with alcohol and then stained with 
a half -saturated aqueous solution of safranin (Schmidt used the Grubler- 

47 See Stiles and Hassall, Sixteenth Report of the Bureau of Animal Industry, 1899. 

48 Arch, of Int. Med., April 15, 191 1, vol. vii, p. 561. 

49 Jour. A. M. A., 1919, vol. 73, p. 1210. 



54 CLINICAL DIAGNOSIS 

Biondi stain) . The specimen is then examined against a white background : 
the mucus stains yellow and albumin red. This method has the advantage 
over other somewhat similar tests that when much pus is present and there- 
fore the albumin in the specimen as a whole considerable, one can estimate 
from the color of the background the relative amount of mucus and albumin 
in the serum of the sputum. In pneumonia the sputum contains much 
more albumin than in other conditions. 

The soluble albumin may be estimated chemically by mixing the sputum with 3% 
acetic acid, shaking it well, allowing it to stand 12 hours, and filtering. The nitrate 
may be tested for albumin with potassium ferrocyanide, or it may be first neutralized, 
sodium chloride added, and the solution tested for albumin by the heat-acid test (see 
page 210). The amount of albumin maybe estimated quantitatively in the Esbach 
tube (see page 216). After filtering out the precipitated albumin the albumoses may 
be precipitated by zinc sulphate or estimated from the amount cf nitrogen present. 
Deutero -albumoses have been found but no peptone (Wanner). Wanner found by far 
the largest quantity of soluble albumin (0.3% to 0.6%) in the sputum of pneumonia; 
little, in that of bronchitis; and the merest trace, if any at all, in that of normal persons. 
The presence of albumin in the sputum means inflammation of the mucous membrane 
of the bronchi, not hypersecretion. 

The quantity of mucin in the sputum may be determined by estimating the amount 
of glucosamin present, of which pure mucin contains 33.6%. To a weighed amount of 
sputum two volumes of alcohol are added, the fluid shaken, filtered through a hardened 
filter paper and the precipitate washed with alcohol. The precipitate is then boiled 
for 3 hours with 10% HC1 in a flask with return cooler. The flask is then quickly 
cooled. Its contents is next made alkaline with NaOH, then acid with acetic acid. The 
biuret-giving bodies are then precipitated with phosphotungstic acid and the amount 
of reducing substance in the filtrate determined with Fehling's solution (see page 176). 
(Glucosamin has the same reducing power as glucose.) From 1 to 3.3% of mucin is 
present in the sputum of chronic bronchitis, a moderate amount in that of pneumonia 
(0.66 to 1.03%), from 0.74 to 0.79% in that of phthisis and none in that of bronchiectasis. 

Wanner 50 considers that in the differential diagnosis between in- 
cipient tuberculosis and chronic bronchitis the presence of a definite trace 
of albumin in the sputum speaks in favor of the former; that the presence 
of considerable albumin indicates pneumonia or pulmonary edema; that 
in the differential diagnosis between pneumonia and infarction of the lung 
the presence of considerable albumin is in favor of the former. Goodman, 51 
after very careful study, concludes that the quantitative estimation of 
albumin is not of very great diagnostic importance, since albumin may or 
may not be present in the sputum of pulmonary tuberculosis and is found 
frequently in the sputum of benign conditions. Its source is generally 
occult blood. 

THE SPUTUM IN DISEASE 

Pulmonary Tuberculosis. — "Pulmonary tuberculosis has no character- 
istic form of sputum " (Brown). "In the earliest stage of tuberculosis the 

50 Deutsch. Arch. f. klin. Med., 1902, lxxv, 347; Fr. Muller, Ztschr. f. Biol., Bd. 52; 
here one finds the best discussion of mucin and its allied bodies in the sputum. 

51 Arch, of Int. Med., August 15, 191 1, vol. viii, p. 163. 



THE SPUTUM: IN DISEASE 55 

sputum will be found negative for tubercle bacilli in from 60 to 75% of 
cases . . . therefore one or even half a dozen negative examinations mean 
nothing. Another very common error is that of assuming the case to be 
one of advanced tuberculosis because the symptoms and physical signs 
seem to clearly indicate it as such . Inasmuch as the diagnosis seems certain 
the sputum is not examined. It is because of this neglect that many 
cases of bronchiectasis, pneumokoniosis, chronic empyema, the mycotic 
infections and even chronic cardiorenal disease, are mistaken for 
tuberculosis " (Landis). 

Cases of chronic fibroid pulmonary tuberculosis may have no, or very 
little (usually purulent), expectoration and the little sputum which some 
cases do raise may for a long time be free of tubercle bacilli. 

In the pulmonic form of acute miliary tuberculosis the patient 
has as a rule little cough and no sputum. When there is sputum it is 
mucopurulent and due to the acute diffuse bronchitis present. Patients 
with the acute bronchopneumonic type of pulmonary tuberculosis expecto- 
rate a rusty or blood-streaked sputum with an occasional hemoptysis. 
Tubercle bacilli if present probably come from an older pulmonary 
lesion. In acute pneumonic tuberculosis the sputum as a rule is identical 
with that of acute lobar pneumonia until the time when the crisis or lysis 
is due, then the appearance of green sputum may suggest the true condition. 
In acute pneumonic tuberculosis with extensive caseous consolidation there 
may for some time be no sputum whatever, but the majority of cases 
expectorate at first a mucoid, then a rusty sputum, which even as early 
as the fourth day may contain Bacillus tuberculosis. Later, in 8 or 10 days, 
the sputum becomes mucopurulent and greenish and then elastic tissue 
may be found. 

Of 15 cases of acute tuberculous lobar pneumonia in the Johns Hopkins Hospital 
Clinic in 4 the sputum was typically rusty ; in the majority it was a mixture of a rusty 
sputum with that of bronchitis ; while 2 had practically no expectoration at all. The 
fresh blood was a marked feature of the sputum of nearly all these cases, while 2 had 
brisk hemorrhages. In the typically rusty sputum very few pus-cells were present, but 
many alveolar epithelial cells and intact red corpuscles. Its green color and tenacious 
consistency were marked in a few cases. After the first week the sputum became more 
mucopurulent and yet still somewhat blood-streaked, later nummular and in 2 cases 
positively foul. In 2 cases, however, the sputum was not at all bloody but muco- 
purulent from the first. 

In those cases in which bronchitis precedes the tuberculous pneumonia there is 
an abrupt change in the sputum. It becomes reduced in amount, tenacious and blood- 
streaked. If a true acute lobar pneumonia develops in the course of chronic tuberculosis 
the sputum is that of lobar pneumonia mixed with that of bronchitis. In 1 case these 
did not mix but formed 2 layers in the cup, the upper mucopurulent and blood- tinged, 
the lower exceedingly tenacious. Later in this case and until death it was a very tena- 
cious greenish pus. In another case elastic tissue was found in the sputum before the 
tubercle bacilli, although repeated examinations for the latter had been made. In one 
case, at first clinically acute lobar pneumonia, the sputum on the third day was markedly 
blood-tinged, contained tubercle bacilli and 2 bronchial casts about 1 mm. in diameter 



56 CLINICAL DIAGNOSIS 

at their larger end. One patient with negative history so far as previous lung trouble 
is concerned had become ill suddenly 2 days before admission. For 9 days his 
sputum was white, sticky, mucopurulent. Then it became green in color and on the 
nineteenth day of his disease he suddenly began to expectorate abundant sputum which 
formed 2 layers, the upper sanguineous and the lower mucopurulent. On that same 
day tubercle bacilli for the first time were found. 

In acute tuberculous bronchopneumonia a hemorrhage is some- 
times the first symptom. The sputum may from the first contain elastic 
tissue and tubercle bacilli. 

In the extensive tuberculous bronchopneumonia which represents the 
terminal stage of this disease and especially in that form which follows an 
acute infectious disease, there may for weeks be no sputum at all while 
other cases will for weeks expectorate a sputum which contains no elastic 
tissue and no tubercle bacilli until necrosis and excavation begin. 

E. N., No. 8288, 20 years old, was admitted June 26, 1919, with extensive broncho- 
pneumonia, presumably tuberculous in character, which followed influenza. The tem- 
perature reached from 103 to 104 each afternoon. She had no cough and no sputum 
whatever till August 17 when she raised just a little which was negative on examination. 
The expectoration of the following day contained both elastic tissue and tubercle bacilli. 
She died September 18, 1919, just 30 days later. Autopsy showed extensive tuberculous 
pneumonia and multiple cavities. 

M. G., No. 9612, aged 22 years, was admitted May 15, 1920, 7 months pregnant, 
with high fever and 'extensive bronchopneumonia with profuse expectoration. This 
was examined almost daily and was negative till June 19, 1920, when numerous bacilli 
suddenly appeared and were present each subsequent day until her return to her home 
soon after. 

In chronic ulcerative tuberculosis the sputum may have almost 
any color and almost any character. It may be (Biermer) mucoid, muco- 
purulent, blood-stained, or almost pure blood. In amount it may vary 
from none to 1 liter in 24 hours and in consistency from that of glue to 
that of water. 

In some cases, especially of early apex tuberculosis with marked physical 
signs and severe cough, there may be no sputum at all, but in the great 
majority of cases there is for weeks a slight morning sputum, although it 
may be necessary to urge the patient to expectorate this. In a few cases 
the onset is with a slight hemorrhage ; in more, the sputum is at first mucus 
containing much myelin and hence is glairy and resembles boiled sago. 
There is nothing distinctive in the gross appearance of such sputum. This 
may be its character for months, but sooner or later one will find in it 
little caseous masses, the first sign of tuberculosis. Later in the course of 
the disease the sputum becomes more profuse and mucopurulent, resembling 
that of chronic bronchitis. As ulceration proceeds it becomes still more 
profuse, mucopurulent or purulent in character and yellow or greenish in 
color. Microscopically, it contains pus-cells, blood and epithelial cells of 
all kinds, including the alveolar epithelial cells filled with myelin, while 



THE SPUTUM: IN DISEASE 57 

if the sputum be hardened en masse and sections cut, giant cells may some- 
times be found. The presence of Streptococcus hemolyticus in these cases 
would seem to be important. 52 

In the course of a case of chronic ulcerative tuberculosis various changes 
may occur. In cases of sudden heart failure there may be a cessation of 
sputum for i or 2 days without apparent injury. "A sudden disappearance 
of the sputum when before it had been abundant, especially in the morning, 
should always put us on our guard. Miliary tuberculosis is occasionally 
ushered in in this manner" (Brown). Again, the sudden appearance of an 
abundant mucoid, more or less frothy sputum may mark the onset of 
miliary tuberculosis (Brown) . 

For diagnosis it is most important to recognize in the sputum the small 
caseous particles, "rice bodies" (corpora oryzoides) since in them one ha? 
the best chance of finding tubercle bacilli and elastic tissue. To find them 
the sputum should be spread out on a dark plate or squeezed between the 
surface of 2 plates of glass (many prefer Petri's dishes which are more 
easily handled and sterilized) and then the whole surface should be scrutin- 
ized with a small hand-lens. They vary from about 0.5 to 1 mm. in diam- 
eter, are of a white opaque color, are more or less lens-shaped, have a bad 
odor, and when picked up with a needle and spread on a slide are found 
to be more brittle and claylike than are, e.g., particles of food (see page 17). 

Elastic Tissue (see page 14). — So important in the diagnosis and prog- 
nosis of cases of pulmonary tuberculosis is the presence of elastic tissue in 
the sputum that the search for it should be methodical and intelligent. 
One may look long and find none, while another choosing particles with 
care may find it easily in a few minutes. Excluding that from other sources 
(see page 17) its presence in • the sputum always means necrosis of lung 
tissue. In tuberculosis one seldom finds the large fragments of tissue found 
in abscess of gangrene of the lung. On the contrary, the disintegration is 
molecular and the elastic tissue is expectorated in the very small, round 
particles mentioned above (also page 6), or in grayish threads, or even as 
single fibers (see page 14) . These fibers come as a rule from the parenchyma 
of the lung and when present in clumps these may preserve the shape of the 
alveoli. Still other fibers come from the bronchi and the blood-vessels. 
In some cases elastic tissue is found early, before there is any suspicion of 
disintegration; in other cases, for instance, in caseous pneumonia, death 
may ensue before any is expectorated. In the majority of cases, however, 
its presence continues as long as ulceration continues and its amount in 
the sputum is a good index of the rapidity of this process. 

Sputum from a Cavity. — Some of the older writers (as Winkel) have 
considered that the odor of the breath may help in the diagnosis of a cavity, 
since in certain cases the sputum in the cup is odorless, while the breath 
of the patient is most offensive (see page 5). The explanation would seem 

52 Hayes, Am. Rev. of Tuberc, 1920, iv, p. 82. 



58 CLINICAL DIAGNOSIS 

to be that warm sputum stagnating in a cavity may have a heavy, penetrat- 
ing, sweetish odor which it imparts to the breath, but not so when cold. 

During cavity excavation the sputum is mucopurulent and often is 
expelled in masses which flatten in the cup into coin-shaped clumps, the 
so-called "nummuli." The best examples of this nummular sputum are 
seen in cases of caseous pneumonia with cavity formation. The nummuli 
are green or dirty grayish green in color and consist chiefly of pus. They 
do not coalesce in the cup. They sink at once in water. Their odor is not 
always offensive. In them can be seen little dots, even as small as a millet 
seed, which consist of granular detritus, black pigment, elastic tissue and a 
few pus-cells. Masses similar to these nummuli may originate also in 
the larger bronchi in non- tuberculous chronic bronchitis. 

When the softening is rapid the expectoration of ioo to 150 c.c. of 
sputum a day is not unusual. The sputum from large cavities is raised 
chiefly in the morning, and often contains blood. If the blood has been 
retained for some time in the cavity it becomes blackish in color. In case 
a tuberculous cavity communicates with a bronchus by a fine opening the 
sputum may form a skein similar to that seen in abscess of the lung (see 
page 78). Large fragments of lung tissue are rarely found in the sputum 
in tuberculosis and yet the connective tissue formation in the walls of a 
healing cavity may be rapid enough to dissect off large particles of the 
necrotic cavity wall which are expectorated. 

While the sputum of cases of tuberculosis with cavity formation usually 
has a sickening, sweetish odor, should secondary infections (other than the 
secondary infections which lead to the cavity formation) lead to bronchiec- 
tasis, gangrene, or putrid bronchitis, the odor of the sputum may be foul; 
but it is remarkable how seldom this occurs. • 

As the cavity clears and becomes lined with connective tissue, the char- 
acter of the sputum changes considerable. We then have the "sputum 
globosum," consisting of balls composed of a conglomerate of mucus, detri- 
tus and pus, of a grayish white color, thick, rounded, and shaggy, some of 
which, but not all, sink in water. 

Hemorrhage. — Hemoptysis occurs at some time or other in from 60 
to 80% of all cases of pulmonary tuberculosis. The amount of blood lost 
on one occasion varies from a drop to several cupfuls. Some patients have 
so many hemorrhages that they constitute the so-called " hemoptysical 
group." In one large group of cases the hemorrhages are early, sometimes 
the first symptom of this disease. These early so-called "inflammatory 
hemorrhages" are slight, apt to recur frequently and are similar to those 
seen in acute lobar pneumonia. Like these they are due to the escape of 
blood-cells by diapedesis or to superficial erosions of the mucosa. Later 
in the disease, however, the hemorrhages are of a very different character. 
They then are profuse and sometimes fatal, often occurring without warn- 
ing in a person apparently recovered. These arise from the rupture of 



THE SPUTUM: IN DISEASE 59 

small miliary aneurisms of arteries which cross an old cavity or which are 
exposed in its wall. 

Sinclair 53 believes that in Hawaii at least hemorrhage rarely occurs 
in incipient cases of pulmonary tuberculosis unless the Spirocheta of 
Vincent and Bacillus fusiformis are present; that when these are present 
hemorrhage occurs in 76% and when absent in but 36% of the cases. He 
admits that the presence of these organisms may be merely an indication 
of a mixed infection. 

For the demonstration of and description of Bacillus tuberculosis see 
page 22. 

One negative examination of sputum proves nothing. Some search for 
3 days, others 6 or 7 ; we believe in searching as long as there is any sputum. 
Brown tells of a patient whose sputum contained tubercle bacilli only on 
the twenty-sixth daily test. "It is of doubtful value to put the sputum in 
the thermostat that the bacilli may grow." 

The Prognostic Value of Sputum Examination. 54 — Sputum examination 
gives very little data of prognostic value. The reasons for this are the 
following: It is certain that not all of the tubercle bacilli in any given 
specimen will take the stain; the sputum from old foci may contain very 
few bacilli and that from young foci, however actively they may be forming, 
may contain none at all ; by the occlusion of a bronchus the contents of an 
intense focus may not be expectorated for a time and when this bronchus 
does open, the sputum, formerly negative, may for a few days contain 
vast numbers of tubercle bacilli and then for months few or none; in some 
specimens the organisms are abundant in one portion and. scanty in others ; 
the sputum of some persons with fatal tuberculosis contain no tubercle 
bacilli at all (for instance, in cases of caseous pneumonia and acute miliary 
tuberculosis), while in other cases the bacilli are present even before one 
can elicit any physical signs of this disease; lastly, in severe cases of tuber- 
culosis with acute bronchitis the bronchial secretion will so dilute the spu- 
tum that the bacilli may appear few in number. And yet the following 
points may assist us in prognosis : 

The continued expectoration of large numbers of tubercle bacilli indi- 
cates the presence of a cavity; a sudden increase in the number of these 
bacilli, together with an increase in the number of cellular elements, sug- 
gests cavity formation; a steady decrease in the number of bacilli continuing 
over a considerable period of time would, if the physical signs agree, indicate 
improvement; a case should be called "healed" only after the bacilli have 
been long absent from the sputum. On the other hand, the continued 
presence in the sputum of large numbers of bacilli does not necessarily 
indicate the case is advancing. Fowler's patient, for example, for 14 years 

53 The Am. Rev. of Tuberc., 1920, vol. iv, p. 201. 

54 Brown, Montreal Med. Jour., October, 1901 ; Jour. Amer. Med. Assoc., February 
2i, 1903. The reader is referred to these articles, from which much of the above 
paragraph is quoted. 



60 CLINICAL DIAGNOSIS 

expectorated daily large numbers of tubercle bacilli and yet during this 
period the patient was in fair health and even improved. Trudeau has 
mentioned a similar case extending over a period of 10 years. Such patients, 
needless to say, are the source of the greatest danger to their neighbors 
expectorating as they may from 3 to 4 billion bacilli each day. 

Many have considered the morphology of the bacilli more important 
in judging of prognosis than their numbers. They hold that while in all 
cases both long and short rods may be found, yet a predominance of short 
rods indicates rapid growth, that of long rods a slower growth. Brown 
believes that while in general the morphology of the bacilli gives but little 
aid, a predominance of short rods does indicate a rather active process. 
Others assert that the arrangement of the bacilli is important; that bacilli 
in clumps and parallel groups indicate a lively growth, and that groups 
of short bacilli mean a bad prognosis. Bacilli which stain deeply are con- 
sidered to be especially virulent. 

There certainly is truth in all of these beliefs, yet there are so many 
exceptions to each that they should not be taken too seriously. 

The question is often asked, Is the discovery of 1 red bacillus in a prop- 
erly stained sputum important ? It would only be suggestive (see page 23). 
A positive report should always be confirmed. On the other hand, the fail- 
ure to find bacilli in the sputum does not necessarily exclude tuberculosis. 

A fairly accurate estimation of the number of bacilli in the sputum is 
possible 55 but for clinical purposes it is not worth the considerable trouble 
it involves. 

Acute Lobar Pneumonia : Croupous Pneumonia. — Some patients at 
first and even during the entire course of a case of acute lobar pneumonia, 
raise no sputum. This is true of some old persons, some alcoholics, of 
infants and young children, very ill patients, and patients with diseases 
of which pneumonia is a complication. This is true often of the terminal 
pneumonias of chronic pulmonary tuberculosis, arteriosclerosis, heart and 
kidney disease, diabetes, etc. But there are also some strong adults, 
especially those with apex pneumonia, who raise no sputum during the 
course of the pneumonia. Some of these patients have more or less dry 
cough, while still others do not even cough during their disease, although 
the absence of sputum in a case otherwise suggesting acute lobar pneumonia 
always should arouse one's suspicions that the consolidation is not due to 
Micrococcus pneumoniae. 

At the onset of a case of acute lobar pneumonia the sputum is as a rule 
quite red, its color due to unchanged red blood-cells, and remarkably trans- 
parent, since these cells are not arranged in rouleaux but are scattered 
singly throughout the mucous mass. Some cases begin with a genuine 
pulmonary hemorrhage. Markedly hemorrhagic sputum, however, should 
arouse the suspicion of tuberculosis since it is very rare in acute lobar 

55 Nuttall, Johns Hopkins Hosp. Bull., Mav, 1891. 



THE SPUTUM: IN DISEASE 61 

pneumonia unless the patients have also heart disease or the case is one of 
post traumatic pneumonia. In still other cases the sputum for even 4 or 5 
days after the onset consists of white mucus, is abundant, and only later 
becomes hemorrhagic. One possible explanation for this is that the clear 
mucus is expectorated while the infection is limited to the bronchi, and that 
the appearance of blood indicates that it has reached the alveoli (Mueller) . 
But whatever the sputum at onset, in from 1 to 3 days from the initial 
chill the great majority of patients with acute lobar pneumonia have the 
typical rusty sputum characteristic of this disease. 

The sputum of few diseases is as distinctive when typical as this. In a 
doubtful case this alone may settle the diagnosis. It is yellowish brown 
in color, the color of iron rust, is homogeneous, glairy, almost transparent, 
and so tenacious and jelly-like that the cup often can be inverted without 
any spilling. Such was the sputum in 7 1% of our cases (counting the rusty 
sputum containing blood) and in 62% of Lord's cases. This rusty color 
is due to a pigment derived from hemoglobin and dissolved in the mucus ; 
microscopically very few intact red blood-cells can be found. This typical 
sputum appears sometimes on the first day of the disease, more often on 
the second, but sometimes later, especially if much bronchitis also is present 
with its mucopurulent sputum. The sputum in pneumonia varies in 
amount from none to from about 150 to 300 c.c. per day. When small in 
amount it dries rapidly in the cup. Its tenacity, which is remarkable since 
it contains little mucin and much water, has been ascribed to nuclein in 
alkaline medium. 

Mixed with the rusty sputum one usually finds fresh blood, often in 
dots or streaks of varying size, while in other cases the sputum for days 
is almost pure blood (see page 82). The extension of the pneumonia to 
another portion of the lung is often indicated by an increase in the amount 
of, or the reappearance of, fresh blood in the sputum. 

While the characteristic sputum of acute lobar pneumonia is rusty in 
color, that of some cases has an orange-yellow, a lemon-yellow, or a grass- 
green color; in fact, all the possible shades which are seen in subcutaneous 
bruises. These colors are due to different oxidation products of hemoglobin 
(Traube). The sputum may appear jaundiced, but this term should never 
be used unless the skin is icteroid for one can practically always get a test 
for bile pigment in pneumonic sputum. 

Microscopically, rusty sputum presents a transparent background in 
which are scattered a few red blood-cells, some intact but the most of them 
swollen and pale and not nearly numerous enough to explain the color; 
many epithelial cells, columnar or pavement; leucocytes; granular cells, 
and oil globules. Chemically this sputum is characterized by an absence 
of alkaline phosphates, an excess of potassium over sodium, an in- 
creased amount of sulphates, of calcium chloride and a large amount of 
soluble proteid. 



62 CLINICAL DIAGNOSIS 

Soon after the crisis the sputum may entirely cease, but more often it 
becomes more abundant, loses its rusty color, becomes mucopurulent, and 
finally is white mucus. " In no other (disease) is the cycle of sputum changes 
so marked or of so great diagnostic value as in this disease" (MacKenzie). 

Of a series of 94 cases, 21 % stated that they had no sputum at the onset of the disease; 
46% had sputum but said that it was not bloody, whereas 33% stated that the first 
sputum noticed was slightly bloody. During the course of the disease 16% of the cases 
had little or almost no sputum; one case was in the hospital 17 days without any expec- 
toration, and other cases about 7 days. In 32% the sputum was typically rusty; in 39% 
rusty and blood-streaked; in 3% very bloody; while in 10% at no time during the disease 
was any blood noted. 

Variations. — If pneumonia develops in a case of chronic bronchitis 
(Traube) the patient usually expectorates a reddish mucopurulent sputum 
which is quite fluid and not at all rusty ; in other cases it is a bloody, mucoid 
pus. In the "hemorrhagic pneumonia" of the aged the sputum instead 
of being rusty is very bloody. If pneumonia develops in a case of chronic 
passive pulmonary congestion due to heart, lung or renal disease, the patient 
expectorates a characteristic "brick-red" sputum which is as thin as that 
of pulmonary edema and very bloody. This is the so-called sputum of 
' ' congestion " or of " serous pneumonia ' ' (Traube) . 

Green sputum in a case of pneumonia running an unusual course always 
should arouse suspicion. If the skin is jaundiced it has no significance and 
if associated with delayed crisis or lysis a perfect resolution may follow. 
But on the other hand it often indicates a serious outcome ; it is sometimes 
the first symptom of abscess of the lung; and, lastly and most important, 
it may be the first hint that the diagnosis of croupous pneumonia was 
incorrect and that the case from the onset was one of tuberculous 
pneumonia. Such were Traube 's cases with green sputum the report of 
which first called attention to this condition. 

In cases of pneumonia which end in necrosis or gangrene of the lung, 
and this is true of 3% of the fatal cases of lobar pneumonia, the sputum 
presents a characteristic series of changes. It first loses its tenacious con- 
sistency and becomes more fluid, its color changes from ' ' rusty " to " coffee, ' ' 
then to ' ' prune-juice, ' ' and later to ' ' chocolate ' ' color. The red blood-cells 
disappear. The odor, at first absent, becomes stale and later decidedly 
fetid. Granular detritus appears and then fragments of necrotic lung 
tissue. In still others of these cases the sputum may be continuously 
" prune- juice " in character, but this is rare. This series of changes in the 
sputum may be sufficient for the diagnosis of pulmonary gangrene even 
before the necrotic fragments have appeared. The reason for these plays 
of colors is not clear since the red blood-cells in some cases have been 
described as well preserved. In about 4% of the fatal cases of croupous 
pneumonia abscess of the lung develops in the consolidated area (see 
page 79). 



THE SPUTUM: IN DISEASE 



63 



Prune-juice sputum in pneumonia has considerable clinical importance. 
It may indicate merely a severe type of the pneumonia ; in still other cases 
it indicates an asthenic type; in others, and in these it has no sinister sig- 
nificance, it merely signifies the beginning of resolution; but it may, as 
mentioned above, warn us of the onset of pulmonary gangrene and, finally, 
its appearance replacing rusty sputum particularly in old persons may 
indicate a developing edema of the lungs. 




Fig. 25. — Fibrin cast from a case of double pneumonia. Natural size. The patient 
was a man 65 years of age. The cast was expectorated on the sixth day of a double 
pneumonia, followed by hemorrhage. Death on the seventh day. 

Fibrin coagula (see page 9) are often present in the sputum of pneu- 
monia, especially in cases of the massive type. They appear at first glance 
as unformed masses, but if shaken out in water are found to be beautiful 
branching fibrin casts of the bronchi, of varying size, the largest with hollow 
branches. Some have clots and small drops of blood in the lumen, especi- 
ally at the bifurcations of the branches. One cast pictured in natural size 
as Fig. 25, seemed to be from 2 entire lobes. Curschmann spirals, and, 
in fact, every other constituent of the sputum of asthma, may be found in 



64 CLINICAL DIAGNOSIS 

that of pneumonia. This was illustrated in Vierordt's case 56 of typical 
pneumonia but with intense general bronchitis. The sputum was bloody 
on the fourth and fifth days and contained many fibrin coagula. These 
were particularly beautiful on the seventh day, on which day spirals were 
also found. Resolution began on this day. From that time on were found 
beautiful Curschmann spirals but no Charcot-Leyden crystals. 

Micrococcus pneumoniae (seepage 32) can as a rule be demonstrated 
in the sputum of cases of acute lobar pneumonia. Since, however, this 
group of organisms is found in the mouths of about 50% of normal persons 
(although its presence there is better determined by animal inoculations 
and by cultures than by smear preparations made from the sputum, see 
page 33), and since it is often found in small numbers in the sputum of 
patients suffering with various other lung diseases, its presence is not 
proof of a pneumococcus infection. On the other hand, its absence from 
the sputum of many cases of acute lobar pneumonia is not to be wondered 
at since at autopsy these organisms are found chiefly at the advancing edge 
of the involved area. 

The term Micrococcus pneumoniae is now used of a group of organisms 
and lobar pneumonia is no longer thought of as due to those of this group 
only. Cole found that 14 of the 237 cases he studied were due to the follow- 
ing organisms: Bacillus influenzae (5), streptococcus (3), Streptococcus 
mucosus (1), staphylococcus (2), Bacillus of Friedlander (2) and one to a 
mixed infection of streptococcus, staphylococcus and influenza bacillus. 
Hastings and Boehm 57 found in the blood and in the sputum of many cases 
of typical lobar pneumonia organisms other than pneumococci; Strepto- 
coccus hemolysans and Streptococcus mucosus capsulatus were found on 
culture in the sputum and Micrococcus pneumoniae in but 24 of the 44 cases 
of pneumonia they studied. The work of Cole and his co-workers (see 
page 33) is however the most important recent contribution to this subject. 

Lobar Pneumonia Due to Friedlander's Bacillus. — Friedlander's 
bacillus is found frequently in mixed infections of the respiratory tract but 
sometimes, in about 4% of the cases, would seem to be the cause of a lobar 
pneumonia, which at first was probably lobular; although even in these 
cases it is often considered a secondary invader. This pneumonia is 
followed by abscess formation and necrosis of lung parenchyma. It runs 
a very rapid fatal course. 

In the cases of acute lobar pneumonia apparently due to this organism 
alone the sputum was very mucoid in character and blood-streaked, rusty, 
or truly hemorrhagic. Microscopically it contained many of these bacilli 
and but few cells. While one may find these organisms in the sputum in 
great numbers the only proof that they are the etiological agent would be a 
pure culture from material obtained by lung puncture, or from blood cultures. 

56 Berl. klin. Wochenschr., July 16, 1883. 

57 Jour. Exp. Med., March 1, 1913, vol. xvii, p. 239. 



THE SPUTUM: IN DISEASE 65 

The pneumonia occurring in typhoid fever is often of the true lobar type, 
especially that which occurs at the onset, the cases of " pneumo typhoid 
fever." The sputum of these patients is sometimes typically rusty and 
may contain Bacillus typhosus. A lobar pneumonia may develop during 
the course of typhoid fever (although bronchopneumonia is the more com- 
mon form). These patients may raise no sputum whatever. It may 
develop also during convalescence. 

In the subacute indurative pneumonia the sputum is usually abundant 
and may contain blood but is seldom rusty. There is a decided tendency 
for it later to become fetid. In chronic interstitial pneumonia the cough 
is often paroxysmal and the expectoration is as a rule copious, of a muco- 
purulent, or a seropurulent nature and sometimes fetid. Hemorrhage 
occurs in about one-half of the cases. 

These forms of chronic pneumonia are often complicated by bron- 
chiectasis, pulmonary abscess, bronchopneumonia or gangrene and these 
conditions may determine the character of the sputum. 

Bronchopneumonia. — Bronchopneumonia may be due to a variety of 
organisms and occurs as a complication in many acute infectious diseases. 
It is a not infrequent complication of typhoid fever. It .is very common 
in severe smallpox and is the rule in severe measles, diphtheria, whooping- 
cough and pulmonary influenza. It is the most common complication of 
typhus fever, in which disease it may result in gangrene, and is common in 
epidemic cerebrospinal meningitis. It is found in acute rheumatic fever, 
rarely in scarlet fever, and is the lesion in pneiimonic plague. Under this 
title are included also hypostatic pneumonia and aspiration pneumonia. 
Children with bronchopneumonia often have no cough, more often raise 
no sputum; and this is true of some adults. The expectoration arises in 
both bronchi and alveoli and so is a mixtiire of rusty and mucopurulent 
sputum. Sometimes the transition from a bronchitis to a bronchopneu- 
monia may be suspected from the changes in the sputum since it becomes 
less in amount, more viscid, more difficult to expel. It may be streaked 
with blood but it is practically never typically rusty. 

In Plague pneumonia the consolidation is lobular and the sputum con- 
tains multitudes of Bacillus pestis. The expectoration may be scanty or 
abundant and when typical is liquid, not tenacious, frothy and bloody. 

Influenza. — Pfeiffer described the characteristic sputum of influenza 
as greenish-yellow in color and expectorated in coin-shaped lumps. Other 
cases, he said expectorated a dark red, bloody sputum. For years the terms 
"influenza" and "grippe" have been interchangeably and carelessly used. 
According to some, grippe is the term which designates all epidemic colds 
with sudden onset and severe prostrating symptoms, whatever the bacteria 
involved, while "influenza" is the term reserved for any condition from 
which Bacillus influenzas (see page 34) of Pfeiffer can be isolated. During 
the pandemic of 1Q18-20 the term "flu" gained considerable popularity 
5 



66 CLINICAL DIAGNOSIS 

and although at first an objectionable abbreviation yet may be the correct 
name of this epidemic since it has been widely used and of this epidemic 
only, while the use of the term influenza would imply that we consider this 
disease the same as the pandemic of 1889 and 1892 which probably is, 
but may not be, the case. This pandemic certainly was due to an, as yet, 
undiscovered organism. 

At this point we would state the belief, which we accept, that the 
"flu" itself, a very contagious disease with leucopenia among its manifes- 
tations and due to an unknown invader, prepares the body for secondary 
infections, and perhaps even for a third or fourth "crop" of organisms, 
which are responsible for the marked and serious manifestations of the 
disease, and that it is this disease together with all its complications and 
sequelae which has been called "influenza" or grippe. 58 

Belonging to the primary infection is a pharyngitis and bronchitis with 
a sputum devoid of characteristic features (except that when introduced 
into the throat of normal individuals does not communicate the disease) 
while the bronchopneumonia which often follows is in many cases (especially 
in the cyanotic cases with low diastatic blood-pressure) characterized by a 
very fluid sputum which consists largely of pure, bright red blood. In many 
other cases, however, the sputum of the pneumonia, usually lobular but 
sometimes lobar, is quite similar to other cases of these conditions. 

These secondary invaders are, Bacillus influenzae and the various strep- 
tococci, earlier the non-hemolytic and later the hemolytic varieties and the 
various pneumococci including Streptococcus mucosus et al. Bacillus 
influenzae of Pfeiffer (1892) has attracted so much attention, for years 
considered the sole causative agent of influenza and now recognized as a 
very important as well as ubiquitous organism, that we describe it at 
length on page 34. 

Whooping-cough. — During the catarrhal stage the cough is, as a rule, 
dry. A little later the sputum is that of bronchitis and presents no especial 
features. During the paroxysmal stage the sputum is expectorated in very 
small amounts each time and yet in the aggregate its amount is consider- 
able. Bordet's bacillus, described on page 37, is now generally accepted 
as the cause of this disease. 

Glanders of the Lung. — In case the glanders infection extends from the 
nose to the bronchi and there excites inflammation, a severe cough accom- 
panied by a profuse purulent expectoration is the result. 

Asthma. — In acute bronchial asthma the sputum may be quite charac- 
teristic. During the paroxysm itself there is often no sputum but expecto- 
ration begins while the paroxysm is subsiding, or, as the patient describes 
it, is "breaking," and brings much relief. But in more typical cases the 
patients expectorate during the paroxysm a tenacious, clear sputum con- 
taining thick, glairy, mucus balls, the so-called "perles of Laennec," which 

5S See Lucke, Wight and Kline, Arch, of Int. Med., 1919, vol. xxiv, 154. 



THE SPUTUM: IN DISEASE 67 

swim in a thin, clear, frothy mucus containing many coarsely granular 
leucocytes and many alveolar cells with myelin degeneration. In still 
other cases the sputum is less characteristic and consists of greenish- 
yellow tenacious mucus described b} r the patient as "rubber-like." It is 
common to find traces of blood in the sputum. These perles are pellets of 
a semi-transparent mucus, of a pearl-gray color like boiled tapioca, some of 
which contain mucous moulds of the smaller tubes and others Curschmann's 
spirals. The moulds are small cylindrical or sausage-shaped masses con- 
sisting of thick threads, or plugs, which may be from i to 1.5 cm. long. 
Some branch, some are narrow or straight, while others are spirally twisted. 
These last have the same significance as Curschmann's spirals. The amount 
of sputum at this stage may vary from a trace to 50 c.c. or even half a 
liter a day. 

In 27% of the Johns Hopkins cases slight hemorrhages had occurred 
in at least 1 of the paroxysms. As the attack "breaks" the sputum 
becomes a clear viscid fluid, thinner, frothy, and more abundant, even 
200 c.c. in 24 hours, and in it float mucopurulent masses, often spirals, 
and in 1 of our cases a true bronchial mucous cast about 1% inches long 
containing many coarsely granular cells. 

During the next 2 or 3 days the character of the sputum changes much. 
It is often small in amount and mucopurulent, mixed with, however, some 
clear frothy fluid. As a rule, no Curschmann's spirals are then found, 
although in 1 case in which they had been abundant they were particularly 
beautiful. Fibrin casts of the bronchi sometimes accompany and may 
outnumber the spirals. Sometimes a branch of a cast ends as the central 
fiber of a typical spiral. With the spirals one also finds in the sputum many 
coarsely granular leucocytes, sometimes Mastzellen, often Charcot-Leyden 
crystals and calcium oxalate crystals. 

The sputum usually ceases as soon as the attack is well over, but it 
may continue, even 100 c.c. per day, or recur at intervals separated by 
sputum-free periods. 

Curschmann's Spirals. — These beautiful structures (see page 7) proba- 
bly appear at some time or other in the sputum of every case of true 
bronchial asthma, but certainly not with every paroxysm. We have in 
mind a man whose sputum furnished the students of several years ago 
with an abundance of beautiful spirals, but during the past 15 years 
although admitted to the ward 14 times during acute attacks of asthma, 
only once was a spiral found. While these spirals may appear at any part 
of the paroxysm they are most numerous just at the end, in the clear mucous 
sputum, and disappear after the sputum becomes mucopurulent. 

Charcot-Leyden Crystals. — Charcot-Leyden crystals (see page 18), found 
wherever eosinophile cells are numerous, are present in abundance in the 
sputum during attacks of asthma. They may occur in groups visible even 
to the naked eye as specks of a greenish-yellow color, which color they 



68 CLINICAL DIAGNOSIS 

may give to the spirals. Their size varies so much that their presence 
can be excluded only after a search with the oil immersion lens. Their 
size and number in the sputum increase as the attack of asthma 
continues, but they often can be found only after the sputum has stood 
in a thermostat. These are much more constant in the sputum of asthma 
than the spirals and do not necessarily indicate the presence of an 
eosinophilia of the blood. 

Alveolar epithelial cells laden with golden-yellow pigment, similar to 
the Hertzfehlerzellen of chronic passive pulmonary congestion, may appear 
in large numbers in definite masses in the sputum in asthma. They may 
fill a considerable part of the mantle of a spiral. 

Certain cases would seem to represent a transitional stage between 
asthma and fibrinous bronchitis, since their sputum contains spirals, 
Charcot-Leyden crystals and eosinophile cells, but also casts of the smaller 
bronchi, the tips of whose branches may be directly continuous with the 
central fiber of a true spiral. In one case of Dr. Osier's 59 already men- 
tioned, these casts were i to 3 cm. long. 

Acute Bronchitis. — Acute bronchitis is a diagnosis frequently and yet 
often erroneously made. It should be used of an acute infection of the 
mucosa of the bronchial trees and yet in most of the cases thus named the 
inflammation seldom extends far down the trachea while in those with a 
definite acute bronchitis this often is overlooked since it is a minor feature 
of a general systemic infection. Another reason for confusion is that with 
an extensive acute bronchitis there probably always is some definite bron- 
chopneumonia. Acute bronchitis is common as a manifestation of the 
infection of influenza, measles, typhoid fever, meningitis, whooping-cough, 
syphilis, typhus fever, smallpox, etc., and in these conditions a broncho- 
pneumonia also is sometimes present. In these cases the bronchitis may 
be due to the organism of the general infection but more often it is 'a com- 
plication due to one of the streptococcus or pneumococcus groups whose 
presence is considered an illustration of symbiosis. Among the organisms 
found in the bronchial secretion are: Bacillus influenzae, Micrococcus 
pneumonias, members of the staphylococcus group and streptococcus 
groups; Micrococcus catarrhalis, as well as the organisms of the specific 
infections mentioned above, Bacillus typhosus, Diplococcus meningitidis, etc. 

In simple acute bronchitis there is often no sputum at all, but if present 
that at the onset is very scanty, frothy, transparent, tenacious, very hard 
to expectorate, the "sputum crudem," of older writers. It consists of 
almost pure mucin, enclosing a few leucocytes, red blood-cells and a few 
bronchial epithelial cells, some ciliated, some even with the cilia in motion. 
The few mononuclear leucocytes present, the so-called "mucous corpus- 
cles," are perhaps derived from the lymphatic masses along the respiratory 
tract. In some cases the sputum contains so many alveolar epithelial cells 

59 See Bettmann, Amer. Jour. Med. Sci., February, 1902. 



THE SPUTUM: IN DISEASE 69 

that the condition has been named ' ' desquamatory bronchial catarrh." 
Myelin drops are found, but only the simpler forms and they are not very 
numerous. Such sputum is due to an hypersecretion of the mucous glands, 
together with the desquamation of a few epithelial cells. While the sputum 
may continue of this character during the whole course of the acute bron- 
chitis, yet as a rule it later becomes mucopurulent. In some of our cases 
it was 2 weeks before very much pus appeared. In others it is at the 
onset more watery and so is termed ' ' seromucous " sputum (Biermer). 
In some cases the sputum is quite bloody at the onset of the attack. This 
was true in 33% of our cases. 

After the first 2 days or more the cough usually "loosens," the sputum 
increases in amount, becomes less viscid, less tenacious, and may resemble 
the white of an egg, since it is frothy and shows whitish streaks. Sometimes 
it is blood-streaked. 

The sputum later becomes mucopurulent. It contains all of the ele- 
ments mentioned above, but with the pus-cells very much increased. These 
may be uniformly distributed and give the sputum a uniform yellow color, 
or they may be present in islands. There are still many epithelial cells 
present but these have lost their shape and their cilia, are now round and 
often fatty. Such sputum was formerly called "sputum coctum." 

In a typical case the sputum next becomes almost pure pus which pours 
from the inflamed and partially denuded mucosa. It is opaque yellow or a 
yellowish-green and is often expectorated in masses. The amount as a 
rule varies from 100 to 200 c.c. in 24 hours, and most in the morning. 
Microscopically the leucocytes are nearly all polymorphonuclears although 
a certain number are mononuclears. No cylindrical cells can now be found. 
Alveolar epithelial cells, some containing pigment and some fat granules, 
may be found if searched for. It contains also much mucus, much myelin 
and fat globules often in masses which in shape suggest cells. In certain 
cases the sputum contains a surprising amount of fat, some in cells, some 
in free droplets but more in the above-mentioned masses of droplets. In 
other cases very little fat is found. The reason for this difference is not 
known (Hoffmann). 

As the case improves the sputum becomes more abundant, more puru- 
lent and less tenacious. It then, as improvement continues, diminishes 
progressively in amount and finally ceases. 

The above is the sequence in a quite typical case. The following vari- 
ations however are met with. In 13% of our cases, the diagnosis of acute 
bronchitis was made because of the physical signs and no sputum was at 
any time obtained. In other cases it was so tenacious that it could scarcely 
be expectorated, the patient often vomiting in the attempt. In some cases 
it was mucopurulent and fairly abundant from the onset but these cases 
probably were acute exacerbations of a slight chronic bronchitis, since 
over 50% of them stated they had been subject to coughs and colds, while 



70 CLINICAL DIAGNOSIS 

that history was obtained from fewer than 20% of those whose illness 
began with scanty expectoration. 

In about 35% of our cases whatever sputum there was was viscid, very 
tenacious and scanty and was followed by a period with a dry cough. In 
about 10% the sputum at the end of the attack consisted of a watery serum 
in which floated islands of mucus and pus about 1 cm. in diameter, which 
settled to the bottom of the cup. Other cases were interesting in that the 
sputum at the end of the attack was similar to that of the beginning, i.e., 
was a pure mucus. 

In the so-called capillary bronchitis, i.e., an acute bronchitis of the 
smaller bronchi, the cough is frequent, often paroxysmal and at first dry. 
There may be no sputum throughout the entire course of the disease or it 
may be scanty and expectorated with great difficulty. In these cases a 
diminution in its viscosity is a sign of improvement. 

The acute bronchitis of early typhoid fever deserves mention since it 
is so constant as to give rise to the adage, " If no bronchitis, it is not typhoid 
fever." The sputum usually is mucopurulent and contains Bacillus typho- 
sus in pure culture or with Micrococcus pneumoniae, one of the streptococci, 
or Bacillus influenzas. 

In pulmonary anthrax, or wool sorters' disease, the condition is usually 
one of bronchitis although pneumonia may develop. 

Chemical Analysis. — The chemistry of the sputum in acute bronchitis 
is of very slight interest. Of the cases which have been reported by Bam- 
berger, Biermer, and Renk, the water content has varied from 95.62 to 
98.3%; the organic substances from 1.17 to 3.7% and the inorganic salts 
from 0.457 to 0.76%. 

Chronic Bronchitis. — Under the heading chronic bronchitis may be 
included all cases of bronchitis from the simple subacute type of a cough 
which has merely "held on" to those cases which give a history of cough 
with expectoration extending over 25 or more years. These long standing 
•cases are usually kept alive by focal infections in the nose, tonsils or mouth. 
The subacute cases expectorate for weeks or months a sputum which is 
scanty, tenacious and viscid. Some patients describe their sputum as 
consisting of thick leathery lumps; others, as a white, sticky mucus. Later 
on it usually becomes more abundant and mucopurulent and hence yellower. 
Some patients for weeks expectorate an abundant sputum with a dark 
greenish color and a foul odor, which tends to separate into 3 layers: a 
mucous layer, a brownish-gray serum and a mucopurulent sediment. The 
sputum in these cases gradually diminishes in amount until the patient is 
apparently well, but he certainly is susceptible to other and similar attacks. 

The bacteriological examination of the sputum of these cases usually 
shows a mixed infection with 2 or more organisms, among them Bacillus 
influenzae, Micrococcus catarrhalis, the pyogenic cocci, pneumococci, 
Bacillus mucosus capsulatus, and Streptococcus capsulatus. Of the com- 



THE SPUTUM: IN DISEASE 71 

paratively pure infections that of Bacillus influenzae is apparently the most 
common, but repeated examinations over long periods usually show that one 
group of organisms does not long remain unmixed in the sputum (Lord) . 

Patients admitted to our hospitals for acute bronchitis usually have 
acute exacerbations of a chronic bronchitis. That is, they have had for a 
long time a dry cough which now has given place to one with sputum, or 
they have had a chronic cough with scanty sputum and now expectorate 
an abundant mucopurulent, often blood-streaked sputum. 

During an acute exacerbation of a chronic bronchitis the sputum is 
sometimes scanty, very tenacious and purulent; sometimes abundant, 
mucopurulent and but slightly tenacious; while in other cases, and these 
are perhaps the most common, is abundant, white, frothy, seromucous 
and contains very little pus. In still other cases the sputum will be very 
large in amount but homogeneous and extremely viscid, a single, purulent, 
glutinous jelly-like mass filling the cup. Its odor is sometimes foul, in one 
of our cases almost putrid. The amount may vary from 100 to 200 c.c. 
in 24 hours. Later the sputum increases in amount and contains small 
mucopurulent flakes. Such sputum separates into 2 layers, a serum 
above and a layer of the solid flakes below. In other cases there is a tena- 
cious green mucous layer above and a fluid layer below. 

The most common type of chronic bronchitis is the so-called "winter 
cough " of patients who during the summer are apparently well. This 
bronchitis may recur for 20 winters or more and yet as a rule the cough soon 
becomes continuous throughout the year. Such cases expectorate chiefly 
in the morning, and describe themselves as then "clear " for the day. Some 
of these patients expectorate each morning about an ounce of mucus, 
while others raise thick yellowish masses of sputum. In severe cases the 
cough is paroxysmal and the sputum sticky, frothy, sometimes blood- 
streaked and very hard to raise. In some cases of chronic bronchitis the 
sputum during an acute exacerbation becomes more scanty and more 
tenacious than before rather than more abundant. These patients feel 
better when their cough loosens. 

Dry Catarrh. — The "catarrhe sec" of Laennec is a symptom-complex 
much in dispute, but cases of chronic bronchitis with little or' no sputum 
are not rare. According to English authors, this occurs particularly in 
"gouty" patients. We associate it, however, more with emphysema and 
nryocarditis. Many of these do raise a little which is glutinous and pearly. 

The chronic bronchitis which accompanies emphysema of the lungs is an especially 
common form. For instance, of 100 cases of long-standing bronchitis, in 43 pulmonary 
emphysema was a marked clinical feature. Of 100 cases of pulmonary emphysema 58 
suffered also from bronchitis, and 47 of these from long-standing bronchitis. This 
cough at first recurs each winter, the patient is comparatively free in summer although 
later it is apt to become continuous. Of these patients with chronic bronchitis 11% 
claimed to expectorate no sputum whatever at any time. In most of the cases with 
scanty sputum the expectoration for years occurred only in the morning, and consisted 



72 CLINICAL DIAGNOSIS 

for the most part of a slight amount of bluish-white tough mucus. It may, however, 
be large in amount. One patient, for instance, for years awakened at 5 o'clock each 
morning with a severe paroxysm of coughing and expectorated a large and almost solid 
mass of thick mucus. In other cases the sputum is abundant, even 1 pint a day, 
frothy and whitish in color. In our cases of emphysema with chronic bronchitis and 
admitted during an acute exacerbation of the bronchitis the changes in the sputum due 
to the acute exacerbation varied much. As a rule its amount increased very materially. 
In one case it became putrid. In one-fifth of the cases it was blood-streaked. As 
these cases improved the sputum first became still more abundant, white and frothy, 
and then gradually diminished to the previous condition. 

In the sputum of 2 cases there were a great many eosinophile cells. Some sputa 
contained large amounts of myelin and others large masses of fat globules. 

One patient with chronic bronchitis had had for 10 or 12 years a slight expecto- 
ration, but on admission his sputum was abundant, thin, cloudy, and contained moulds 
of the bronchi, the stem of some 0.5 mm. in diameter, which consisted of mucus, pus 
and alveolar epithelial cells. This sputum also contained much pigmented alveolar 
epithelium, pus-cells and red blood-cells. 

One emphysematous patient with long-standing bronchitis, admitted during a 
rather acute attack with continuous fever, expectorated sputum which was abundant, 
seromucoid, never bloody and which contained during repeated examinations large 
numbers of sarcinae. 

In 1 case of chronic bronchitis with emphysema and "hay fever" the sputum was 
yellowish-green, mucopurulent, slightly blood-tinged and contained branched bronchial 
plugs which consisted of mucus, pus-cells, many eosinophile cells and masses of the 
mycelial threads of some mould. 

In the chronic bronchitis of mitral cardiac disease the characteristic 
sputum contains a large amount of blood, which gives a bright-red or prune- 
juice color. In other cases, particularly of mitral stenosis, it is often stained 
by masses of Hertzfehlerzellen. Other cases, however, expectorate merely 
large amounts of frothy, seromucous pus. 

Bronchorrhea. — If by bronchorrhea one means, with Laennec, a 
chronic idiopathic disease characterized by the expectoration of large 
amounts of watery sputum we may doubt the existence of such a disease. 
But if we mean merely chronic bronchitis with an abundant watery sputum 
the condition is by no means rare. Some have described a ' ' bronchorrhea 
serosa," or "asthma humidum," with abundant, very watery, colorless, 
foamy sputum. Some of these cases are said to have a neurotic basis. 
In the bronchorrhea of chronic bronchitis the patient may expectorate 
about 500 c.c. a day which is as a rule purulent, watery and of a green or a 
yellowish-green color. In cases of "bronchoblennorrhea," with bronchi 
denuded of mucosa and lined by a pyogenic membrane, the sputum con- 
tains very little mucus, is a profuse watery pus which separates easily 
into 3 layers and which may have a very bad, although not a distinctly 
fetid, odor. Sputum somewhat similar is seen also in bronchiectasis and 
perhaps also in cases of putrid bronchitis and lung gangrene. 

Putrid Bronchitis. — In many cases of chronic bronchitis the sputum 
has a disagreeable, almost fetid, odor; but in putrid bronchitis it is truly 
fetid. Fetid sputum occurs also in many cases of bronchiectasis, in gan- 



THE SPUTUM: IN DISEASE 73 

grene of the lung, in abscess, in tuberculosis with large cavities and in 
empyema perforating through the lung. Cases of fetid bronchitis with the 
bronchi not dilated certainly are very rare (Fowler and Godley ) , some even 
deny that they exist (Hoffmann) and claim that these were cases of bron- 
chiectasis, which in time any case of putrid bronchitis would soon become. 
A very few genuine cases of putrid bronchitis have, however, come to 
autopsy (Osier). 

The sputum in putrid bronchitis is profuse, watery, of a dirty ash-gray 
or a brownish color, and with a horrible odor which will fill the whole house. 
Allowed to stand, it separates into an upper layer of frothy air-containing 
mucus, usually small in amount since the mucous membrane is for the most 
part destroyed and from which layer stream downward brownish strands ; 
a middle layer of serum, and the lowest a thick sediment of epithelial cells, 
fatty cells, free fat, almost pure pus, all kinds of bacteria and sometimes 
Dittrich's plugs. No elastic tissue fragments of lung are to be found. 

Gangrene may follow in such cases. 

Chemically one finds in the sputum of these cases many of the products 
of the decomposition of proteids; volatile acids, among them butyric and 
valeric; NH 3 , H 2 S, leucin, tyrosin, etc. 

Fibrinous, Croupous, or Plastic Bronchitis. — The acute form of 
fibrinous bronchitis which accompanies certain infectious fevers is men- 
tioned on page 9. Chronic idiopathic fibrinous bronchitis is so rare a 
disease that Bettmann could find but 2 7 cases in the literature of 3 5 years. 
During an attack of fibrinous bronchitis the sputum, for 5 or more days, 
consists merely of abundant mucus and then suddenly there develops a 
severe coughing spell and the patient expectorates a bronchial cast. Blood 
usually accompanies the cast but may precede or follow it. Profuse hem- 
orrhages are rare. The frequency with which casts are expectorated varies 
much. Usually but one is expectorated during several months but some 
patients expectorate 1 every 2 or 3 days while 1 patient expectorated 
3 during the same day. The casts appear in the sputum as formless masses, 
but if shaken out in water are found to be moulds of a bronchial tree. All 
from the same patient are apt to be exactly alike as though all came from 
the same bronchus. Some would seem to represent the bronchial tree of an 
entire lobe. The largest casts are about 10 cm. long, are grayish -white in 
color, contain a great many air bubbles and in about one-third of the cases 
are blood-streaked or contain a blood-clot in their lumen. They consist 
of a fibrin-like material in layers arranged concentrically, the innermost 
of which presents many whorls, since this was the first formed and this is 
much compressed by the layers peripheral to it and formed later. The 
casts are loose in structure, are usually hollow, although some are solid; 
of others the larger branches are hollow and the smaller solid, while in others 
the reverse is true. In the central layer, the oldest, are seen the remains of 
cells, alveolar and bronchial epithelium, leucocytes, red blood-cells and 



74 CLINICAL DIAGNOSIS 

bacteria, but in general these casts contain much air and few cells. Some- 
times the casts contain much fat as does also the sputum. 

Casts are not always products of the epithelial cells of the mucosa. 
There was, for example, no epithelium in that part of the bronchial tree 
from whence the casts came in the case of the above-mentioned patient 
who expectorated 3 in 1 day. Those casts must therefore have been the 
result of direct exudation. 

Bronchial casts were formerly supposed to consist of fibrin, since their 
physical appearance suggests this. Others claim it is mucus, others say 
syntonin or coagulated albumin. This material sometimes takes Weigert's 
fibrin stain, but usually it does not. Liebermeister 60 reviewed this question 
at length as the result of the study of 1 fresh case and of 1 2 museum speci- 
mens. He demonstrated fibrin and mucin in 7 of these 1 3 cases. To demon- 
strate fibrin he used Kockel's method, and thionin for the mucin. 

Charcot-Leyden crystals are commonly present in these casts. In the 
same sputum spirals also sometimes are found. In Vierordt's 61 case there 
were many such casts, and on one occasion a typical Curschmann spiral. 
In Dr. Osier's case, mentioned by Bettmann, the ends of some branches of 
the casts were directfy continuous with the central threads of true spirals. 
Many eosinophile cells also are present, also red blood-cells, hematoidin 
crystals, and lecithin granules. 

Casts similar in appearance appear in the sputum of other conditions: 
in diphtheria, which casts are firm hollow tubes of dense fibrin enclosing 
countless cells; those in pneumonia are mentioned on page 63; and the 
cast from 1 case of heart disease was similar to those of fibrinous bronchitis. 

Syphilis of the Trachea and Bronchi. — In syphilis of the trachea 
and bronchi the cough is at first dry but later is accompanied by mucoid, 
mucopurulent, or purulent sputum which often is blood-stained and which 
sometimes contains also elastic tissue. Hemoptysis has been a prominent 
feature in about one-half of the cases, while about half of the fatal cases 
have died from hemorrhage from ulceration of the luetic lesion of the trachea 
or larger bronchi through into the pulmonary artery, aorta, bronchial 
artery or superior vena cava. 

Bronchiectasis. — The sputum in the saccular form of bronchiectasis 
may be quite characteristic; in the diffuse form it is practically never so. 
The former is marked by 2 features — its large amount and its periodicity. 
That is, the patient occasionally, and usually following some definite change 
of posture, will have a paroxysm of coughing, often severe, and expectorate 
a large amount of sputum. Having emptied the sac of the dilated bronchus 
he may for hours be free from cough and sputum. Since this follows a 
marked change in the position of the body, it most often occurs on rising 
in the morning, but the posture which will liberate the coughing reflex 

60 Deutsch, Arch. f. klin. Med., 1904, Bd. 80, 5, and 6. 

61 Berl. klin. Wchenschr., July 16, 1883. 



THE SPUTUM: IN DISEASE 75 

will depend in a large measure on the position of the sac. This periodic 
feature was marked in but 10 of our 24 cases. The amount of sputum these 
patients have is considerable, as a rule from 750 to 900 c.c. in 24 hours, 
but in 1 of our cases it frequently exceeded 1 liter. Such profuse expectora- 
tion may extend over a considerable period of time. 

Of 23 cases, in 2 the sputum for 24 hours was under 100 c.c. ; in 1 1, from 1 to 300; in 2, 
about 500; while in 7, it exceeded 600 c.c. In general the amount of sputum bears but 
little relation to the duration of the disease. One of our cases, of 26 years' standing, 
expectorated only from 15 to 30 c.c. a day. It bears little relation to the size of the 
cavity, as was shown by 1 of our patients who expectorated more than 1 liter of sputum 
each day and yet at autopsy a few, surprisingly small, cavities were found. It is stated 
that there is a remarkable diminution in amount of sputum as the patient grows weak 
before death. 

The most characteristic sputum in bronchiectasis is grayish or grayish- 
brown in color, fluid, purulent, of a disagreeable odor and separates on 
standing into 3 layers. But this is not the only form. Early, while the 
bronchiectatic cavity is lined by mucous membrane, the sputum is a pure, 
clear mucus, but soon the cavity becomes infected and then the mucosa 
becomes a pyogenic membrane which produces a yellow, purulent fluid 
with a sweetish odor. Sooner or later putrefactive organisms invade the 
cavity and then the sputum becomes of any shade of gray or green, muco- 
purulent, and has a horrible fetid odor. Those of our cases with the worst 
odor were dirty gray in color. There is usually bleeding into the cavity 
and then the color of the sputum may have any shade of red or brown 
according to the chemical changes which the hemoglobin undergoes. 
While, as a rule, the sputum is very fluid and watery, in some cases it is 
thick and viscid, while in other cases it is mucopurulent and contains 
masses suggesting nummuli. In certain cases well illustrated in our 
series, particularly those which improved under treatment, the sputum, 
which was at first profuse and watery, later diminished in amount, became 
mucopurulent and of a less offensive odor. The tendency to form 3 layers 
on standing in a tall glass vessel was marked in 14 cases. These layers are : 
an upper frothy mucous layer, a middle serous, and a lower, always thick, 
granular layer. From the upper layer often hang down through the fluid 
' ' streamers ' ' of mucus laden with pus. This is often spoken of as the second 
layer. Hoffmann and others mention but 2 layers, omitting the upper 
which was absent in 3 of our cases. 

In 4 of our cases there were 4 well-marked layers; the lowest, abundant, of greenish- 
red purulent material; the one above, containing a good deal of blood and hence red or 
brown ; over this a serous layer and on top a frothy mucous layer. In other cases below 
the top layer was a mucopurulent layer of streamers hanging down through the fluid, 
which with a little encouragement would probably all have sunk to the bottom. The 
odor is, in general, bad, but in 2 of our cases it was not at all offensive. In some cases 
it had at first no odor, then became slightly offensive, while later, after the putrefactive 
changes had set in, it was fetid. These changes are due to secondary infections of the 
cavity. In 10 of our cases the odor was heavy and sweet, while in 10 others it was at 



75 CLINICAL DIAGNOSIS 

some time very fetid. This fetid odor is not exactly the same as that present in gangrene 
and has been described as " pseudogangrenous " in character, the odor of rotten cabbage, 
or garlic. This odor will often diminish after long-continued treatment with creosote 
inhalations or intratracheal injections. Indeed a patient admitted with extremely fetid 
sputum may leave the hospital with expectoration much reduced in amount and not 
at all offensive in odor. The breath of the patients is sometimes worse than their sputum. 
The disagreeable odor is largely due to H 2 S, NH 3 , and various volatile acids, among 
which are acetic, butyric, and formic acids. 

Hemorrhages into the bronchiectatic cavities are common, occurring 
in even 50% of all cases (in 17 of our 24 cases). The hemorrhage is slight 
as a rule (8 of our cases) but sometimes (6 cases) is considerable in amount, 
while in 3 it was extreme. In some cases it has been fatal. 

One of our cases was admitted to the hospital 14 times, 5 because of extreme hemor- 
rhages which threatened his life. During one of these admissions he had, in a very few 
days, 6 large and several smaller hemorrhages, which reduced his blood count rapidly 
from about normal to 1,090,000 red blood-cells and the hemoglobin to 20%. Another 
case in 1 day and in about 10 minutes lost 1700 c.c. of blood and on the following 
day died from a hemorrhage. 

Microscopical Constituents. — As long as a bronchiectatic cavity 
is not infected its contents will consist of mucous and desquamated epi- 
thelium cells. After infection its wall becomes a pyogenic membrane and 
its contents those of an abscess. Later, infection with the organisms of 
putrefaction is the rule. The pus-cells, enormous in numbers, are well 
preserved, fatty, or vacillated. The red blood-cells are unchanged or very 
much altered. Elastic tissue theoretically should not be found and yet it 
was present in 2 of our cases, indicating ulceration of the bronchial walls. 
Fatty acid crystals are numerous especially when the outlet of the cavity 
is small, thus allowing considerable stagnation of its contents. These 
crystals occur in large masses, are very large in size and present a beautiful 
picture. They were abundant in 4 of our cases (see Fig. 8). Cholesterol is 
often present; hematoiden crystals frequently; leucin and tyrosin some- 
times; and Dittrich's plugs, usually. Alveolar epithelial cells are often 
present, some containing pigment and others much myelin and fat. No 
tubercle bacilli are found, but other bacteria are, in great numbers and form 
large zoGglea. Yeasts are found, and, in one of our cases, an aspergillus 
mould. Calcium salts are sometimes deposited in the contents of these 
cavities, giving rise either to a clay -like mass or as in 2 of our cases to a 
bronchiolith. In 1 of these cases, that of a man who during life has 
expectorated several concretions, the pathologist found at autopsy consider- 
able calcareous matter embedded in the walls of a cavity. The bronchioliths 
in his sputum were about the size of split peas. 

In other cases, as in 13 of our series, the sputum is by no means so 
characteristic and resembles that of a chronic or of a fetid bronchitis 

Children with bronchiectasis are apt to swallow and then to vomit 
their sputum. 



THE SPUTUM: IN DISEASE 77 

Gangrene of the Lung. — The most characteristic sputum of patients 
with pulmonary gangrene is profuse in amount, watery, greenish-brown or 
ashy-gray in color, has an extremely fetid odor and separates easily into 
layers. But more commonly it contains blood and so has a color which 
may vary from reddish-brown to brownish-red or even chocolate. In other 
cases it has a uniform dirty-brown color due to masses of hematoidin 
crystals. Its odor usually is the worst of all the sputa and yet in some cases 
of true gangrene of the lung the sputum and the breath have no odor at all. 
In 5 of 12 cases of the Johns Hopkins Hospital the presence of the gangrene 
was not even suspected during life. One case had expectorated merely 
"phlegm." These patients are usually diabetics or insane. In cases of 
pulmonary embolism the infarcted area may become gangrenous and later 
be evacuated through a bronchus. 

It is often difficult to differentiate between gangrene and abscess of the 
lung, since whichever is primary the other is quite sure later to develop. 
The presence of tissue fragments will aid in the differential diagnosis 
between gangrene and long-standing bronchiectasis. Putrid bronchitis 
differs from gangrene only in the absence in the sputum of fragments of 
lung tissue. This sputum separates easily into 3 layers — the upper of 
frothy mucus, the middle of serum and the lowest, always a thick one, 
of pus, tissue detritus, Dittrich's plugs and tissue fragments. From the 
top layer streamers often extend down through the fluid. In other cases, 
the sputum is mucopurulent in nature, while in others it is viscid, lumpy, 
mixed with blood and also very fetid. 

Of the macroscopic constituents of the sputum of cases of pulmonary 
gangrene the fragments of necrotic tissue are the most important and must 
be demonstrated before a positive diagnosis is possible. It is in this disease 
that the largest fragments of lung are expectorated. Some are very 
minute but others are even several centimeters in length. Some are firm 
in texture, sooty in appearance, have ragged outline, while others consist 
of a colorless ground substance full of granular detritus, fat droplets, 
clumps of coal, large fat needles, bacteria and elastic tissue. The other 
constituents of the sputum are those of fetid bronchitis. 

It is said that elastic tissue is rarely if ever found in the sputum of this 
disease and that its absence is due to some ferment which digests it in the 
tissue masses. The chances are, however, that it can always be found if 
searched for. To be of aid in diagnosis the elastic tissue must show by its 
arrangement its origin in the alveolar walls. Alveolar epithelium, often 
pigmented, is easily found ; fatty acid crystals and fat droplets are abundant ; 
cholestrol, leucin, and tyrosin may be demonstrated; masses of bacteria 
and often of leptothrix are conspicuous, while flagellata have been described. 
In a case mentioned by Sahli, in which the infected area was non-odorous, 
large numbers of sarcinas were found. Blood is frequently present and in 
large amounts, the origin of which is the rupture of small vessels, not 



78 CLINICAL DIAGNOSIS 

diapedesis. Fresh blood was present in 5 of our cases, but as a rule the 
hemoglobin is present as methemoglobin and hematin. 

Many observers (e.g., Mayer) in 10 of 58 cases have found acid-resisting 
but not alcohol-fast organisms in the sputum of these cases (see page 30). 

Aspirated Foreign Bodies. — The aspiration of a foreign body immedi- 
ately gives rise to a paroxysm of cough, suffocation, cyanosis and intense 
dyspnea. If the foreign body remains impacted this soon subsides and a 
period of comparative relief may follow which often leads to a false sense 
of security marked only by occasional slight cough and dyspnea on exertion. 
In a few days, however, the manifestation of secondary bronchial infection 
appears, i.e., cough, with more or less abundant sputum which is purulent 
and soon foul-smelling, then the clinical features of pulmonary abscess 
develop which may last for weeks, months, or even years. 

Abscess of the Lungs. — Abscesses of the lung are evidently much more 
common than was believed. Indeed they may follow any operation on an 
infected tissue, even simple tonsillectomy, and may be a sequela of any 
bronchopneumonia due to a pyogenic organism, as the so-called ether 
pneumonia, metastatic pneumonia, etc., as well as a common sequela of 
influenza. They may rupture into a bronchus causing no marked symptoms 
if small, or into the pleural cavity causing empyema. An abscess of the 
lung may be suspected if the patient suddenly expectorates a large amount, 
even several hundred cubic centimeters, of quite pure pus in which are 
fragments of lung tissue. If allowed to stand this sputum will separate 
into several indefinite layers, but layer formation is not definitive since a 
slight shaking will restore the sputum to its previous homogeneity. The 
odor of this sputum is at first faintly sweet like all pus, but if lung gangrene 
develops as it often does, it becomes foul. The lung-tissue fragments are 
very important in diagnosis. In size they vary from about that of a millet 
seed to fragments even 2 inches long. In the larger fragments of lung 
tissue may be demonstrated a framework of elastic tissue, the remains of 
blood-vessels, masses of coal dust, fat crystals, free fat, detritus, hema- 
toidin crystals, amorphous clumps of pigment, and zooglea of cocci. In 
other cases one finds no large fragments. In these the lungs undergo the 
so-called "insensible disintegration" (Ley den). In this sputum one finds 
separate elastic fibers and free cholesterol, fatty acid crystals, free fat, 
lung pigment, detritus, bacteria and hematoidin crystals which may be 
present in large numbers and give to the whole mass of sputum a brown 
color. Older writers have described a sputum the gross appearance of 
which was quite characteristic of lung abscess. If one shakes it out in 
water it would appear like a skein or thread of pus. The explanation of 
this would seem to be that the pus escaped in a thin thread from a large 
cavity slowly through a small opening and later received in the bronchi 
a mucous coating which prevents its coalescence. We have seen no 
such case. 



THE SPUTUM: IN DISEASE 79 

The sputum of a liver abscess perforating through the lung is often char- 
acteristic because of the bitter taste of the bile acids, its so-called "anchovy- 
sauce" appearance, or its ocher-yellow color from bile pigment. In some 
of these cases a lung abscess also develops, in other cases only a simple 
hepaticobronchial fistula. Microscopically bilirubin crystals and much 
elastic tissue will be found and sometimes the amebse themselves. 

In the records of the Johns Hopkins Hospital there were 7 such cases. In 3 the 
sputum was abundant, even over a liter in 24 hours. In some the expectoration was 
paroxysmal, even a quart at a time. The odor was mildly offensive in 2 and markedly 
so in 2 others. In six of these cases the sputum presented the typical "anchovy-sauce" 
appearance; that is, it was of a rusty brownish-red color and frothy. In 4 cases it 
was blood-streaked and in 2 purulent. Microscopically were found the ordinary 
elements of sputum: pus, red blood-cells and alveolar epithelial cells, and, in addition, 
fat crystals and the crystals or needles of hematoidin (bilirubin) which were a marked 
feature in 2 cases. Elastic tissue was found in considerable amounts in 5 cases. 
In 2 cases the liver cells, it was thought, could be recognized. The living active 
amebae were found in the sputum of 5 cases, on 1 even before they were found in the 
stools. The sputum which contained the ameba also contained much elastic tissue. 
If the sputum be preserved in the thermostat the ameba will remain alive and motile 
for even a whole day. 

The influenza epidemics were followed by many cases of lung abscess, 
the logical results of the areas of interstitial pneumonia. We believed 
that many of the cases of empyema were due to the rupture of 1 of these 
abscesses into the pleural cavity. 

E. S., No. 7554, a boy aged 16, was admitted Dec. 15, 19 18, following an attack of 
influenza which began Nov. 21, 1918. On December 16 he suddenly coughed up 300 c.c. 
of thin, purulent sputum with a sweetish yet fetid odor. Smaller amounts followed. 
On standing this sputum separated into 3 layers, the upper of mucus, the middle a thin 
turbid grayish liquid and the lower which was thick, ropy and salmon colored. Four days 
later 1600 c.c. of foul smelling thick pus were aspirated from the right chest. This did 
not separate into layers on standing and contained many streptococci. The left lung and 
pleural cavity were quite normal throughout. He was discharged well March 28, 19 19. 

Abscess of the Lung Following Acute Lobar Pneumonia. — In 
3 of 6 cases of fatal pneumonia with pulmonary abscess (found at autopsy) 
there had been no clinical features which suggested this complication. In 
these cases the abscesses were small and multiple and there had been little 
or no sputum. In 1 of these cases the only change noted in the sputum 
was that the viscid, tenacious, blood-streaked expectoration became less 
tenacious. In 1 case the sputum, which had been small in amount, very 
tenacious and blood-tinged, suddenly became very dark brownish-black 
in color and mucopurulent. It then became greenish in color and small 
in amount. Then all expectoration stopped, soon to begin again as a 
mucopurulent sputum. It then became very green and scanty and later 
increased much in amount, was very thick, very purulent and of a sour 
odor. It then became thinner, more watery, blood-stained and contained 
elastic tissue. Then it decreased in amount, became mucopurulent, and 



80 CLINICAL DIAGNOSIS 

finally, with the recovery of the case, ceased. In i case large numbers of 
trichomonads were found in the sputum. 

Three cases of post-operative pulmonary abscess were followed clinically. 
One patient who was admitted with a paroxysmal cough suddenly expec- 
torated a large amount of foul-smelling sputum. The odor later became 
less offensive. It contained pus and fatty acid crystals. This sputum con- 
tained large fragments, even 5 by 3 cm. in size, evidently of tissue but so 
decomposed that their structure could not be determined. The sputum 
then became less profuse, then mucopurulent and the patient recovered. 
In another case the sputum was very foul-smelling and contained much 
fat, while in another case it was large in amount, purulent, foul-smelling 
and blood-streaked. Of 2 other cases not recognized during life 1 had 
showed a sudden increase in the amount of sputum, while in the other an 
abundant, blood-streaked, brownish sputum of no especial odor suddenly 
increased in amount, became dirty, frothy, foul-smelling, slightly blood- 
streaked and separated easily into 3 layers. At autopsy a large abscess 
cavity was found. 

Perforating Empyema. — The sputum of cases of pleural empyema 
which perforates through the lung resembles that of abscess of the lung, 
with the exception, however, that that of the former contains less elastic 
tissue and practically no tissue fragments. It contains many hematoidin, 
as well as other, crystals. Its odor is at first that of pus, in some cases 
described as resembling old cheese, but later it becomes vile because of 
the secondary infections which commonly follow. In case the pleural 
fluid escapes slowly through a small pleurobronchial fistula, it is said that 
a skein of a thread of pus may form similar to that described on page 
78. When the opening is large the pus will escape rapidly, yet without 
causing pneumothorax. Allowed to stand, this sputum separates into 
3 layers, the upper of mucus, the middle of the pus-serum and the lowest 
of pus-cells. 

Perforating Serous Pleurisy. — It is exceedingly rare for a serous pleural 
effusion to rupture through the lung. The sputum in such a case is like 
that of edema of the lungs, but contains more albumin and so becomes quite 
solid on boiling. 

The Serous Sputum of Edema of the Lungs. — Patients with edema 
of the lungs expectorate large amounts of a frothy, cloudy, colorless, or a 
slightly bloody sputum which on standing separates into 3 layers : an 
upper abundant frothy layer, a middle foamy fluid, and a thin lower layer 
of pus together with the elements of the pre-existing sputum. Excepting 
in cases of pneumonia, etc., the sputum is quite pure blood-serum. It is 
frothy since it is so rich in albumin and watery since it contains only a trace 
of mucin. Patients during the last hours of life with this sputum flowing 
in continuous streams from their nostrils and mouth present one of the 
most gruesome sights of the wards. 



THE SPUTUM: IN DISEASE 81 

The Albuminous Expectoration of Thoracentesis. — Among the recent 
important articles on albuminous expectoration are those of Riesman 62 
and of Allen. 63 Terrillon has grouped cases of this condition into 3 classes: 
The first of mild cases with sputum varying in amount from little to 800 
c.c. ; the condition of these patients is always good. The severe cases have 
dyspnea and collapse, and expectorate from 1200 to 1500 c.c. In the grave 
cases the fluid may suddenly gush from the mouth. Some patients are 
drowned in the fluid which they cannot expectorate rapidly enough, while 
others have died before they could expectorate any. 

The expectoration of this albuminous sputum may begin with a par- 
oxysm of coughing during aspiration, or in less than 1 hour after this is 
finished, while in the latest cases it begins 18 hours after the tapping and 
continues for 24 hours. As a rule it lasts from 1 to 2 hours. This sputum 
is rich in albumin, frothy and neutral or faintly alkaline in reaction. To 
test it for albumin the sputum should be diluted and filtered and the filtrate 
tested by heat and nitric acid, by nitric acid alone, or by potassium ferro- 
cyanide. Acetic acid without heat gives a precipitate of mucin. This 
sputum also contains urea, hemaglobin, urobilin and the various salts of 
blood-serum. The amount expectorated averages from 200 to 900 c.c. 
while even 2 liters have been expectorated. On standing 3 layers separate — 
the upper, whitish and frothy, the middle, opalescent and yellowish or 
greenish and the lower, more viscid, contains a few whitish flocculi and 
sometimes slight traces of blood, but rarely much. In Riesman 's case 
there was no lower layer, the specific gravity was 1.018, the fluid became 
solid on heating. The total solids were 5.84%. In Allen's case the expec- 
toration began in half an hour after 3100 c.c. of pleural fluid had been 
removed and lasted 4 hours. It measured about 1 liter in amount, was 
frothy, pale green in color, with a muddy sediment. Microscopically were 
found flat epithelial cells, a few leucocytes, red-blood cells and many 
bacteria. Other such sputa have had quite different composition. Some 
resembled pleural exudates. 

The cause of albuminous expectoration has been in much dispute. The 
majority of writers think that it is due to an acute edema of the lungs and 
is the result of too rapid or too complete thoracentesis. We would call 
attention, however, to certain cases which occurred during thoracentesis 
and which were followed by pneumothorax. In these cases the pleural 
exudate itself may have been evacuated through the mouth. 

Hemoptysis. — For the causes of pulmonary hemorrhage we quote 
both Osier and Lord. 

Hemoptysis may occur: (1) In young healthy persons, without known 
cause and without subsequent symptoms. (2) As the first symptoms of 
pulmonary tuberculosis. This explains 7 7 .6% of the cases of sudden hemor- 

62 Amer. Jour. Med. Sci., April, 1902, p. 620. 

63 Johns Hopkins Hosp. Bull., January, 1903. 

6 



82 CLINICAL DIAGNOSIS 

rhage in healthy voting men and is due to mucous erosions and to the dia- 
pedesis of red cells through the congested mucosa. Sudden hemorrhage 
was noted in 900, or 0.045%, of the soldiers of the Prussian army studied 
with this point in view, all apparently healthy young men. In 480, or 54%, 
there was at the time no apparent cause. Of these 480,417, or 86%, were 
probably tuberculous and 221, or 46%, quite certainly so. (3) In an ad- 
vanced case of pulmonary tuberculosis or in a cured case with cavity forma- 
tion, due to the rupture of a miliary aneurism on a branch of the pulmonary 
artery exposed by cavity formation. (4) In other diseases of the lungs, 
and this list includes practically all pulmonary diseases. Among these are 
pneumonia at the onset, "bloody bronchitis," cancer, gangrene, abscess, 
bronchiectasis, tumors, cysts and actinomycosis. (5) Heart disease, espe- 
cially mitral. As a rule these hemorrhages are slight, yet they may be 
profuse and may recur for years. (6) Vascular degeneration, the result of 
increased pulmonary tension, seen in emphysema and arteriosclerosis. 
(7) In ulcerations of the larynx, trachea, and bronchi, in which cases it 
may be profuse and rapidly fatal. (8) In aneurisms of the large vessels 
of the chest it is sometimes sudden and fatal; in other cases the so-called 
"weeping" may persist for weeks, or the pressure of the aneurism may 
cause a bleeding erosion of the tracheal mucosa. (9) An extremely rare 
form of vicarious hemorrhage due to interrupted menstruation. (10) In 
rheumatism. (11) Malignant fevers, the so-called hemorrhagic type. 
(12) Purpura hemorrhagica and various other blood diseases, among which 
are hemophilia, leukemia and scurvy. (13) Distomatosis (Westermanii) . 
The amount of the blood expectorated in any of the above conditions 
may vary from a drop or a few small clots to a quart or more. In general 
it has a bright red color even when of venous origin since it is aerated in 
the lungs and is frothy from its admixture with air. The blood may clot 
in the bronchi and casts of these be expectorated. In gastric hemorrhage 
the blood is as a rule dark because of the acid gastric juice, not frothy, 
partly coagulated and is raised by vomiting. If, however, the stomach is 
empty when a large gastric artery is opened this blood may be bright, 
while a profuse hemorrhage from a pulmonary artery may be dark and not 
frothy. Such points would be easy enough to determine were the doctor 
the observer, but a diagnosis from the history given is often difficult. 
Paroxysms of severe coughing often cause vomiting, while the coughed 
blood may be swallowed and vomited, or a little vomited blood may be 
inspired and set up a paroxysm of coughing. A history of previous lung 
or stomach trouble is important in diagnosis, also to follow the sputum 
which for some days after a pulmonary hemorrhage will be blood-tinged, 
and the stools which will contain blood for a few days after a hemorrhage 
from the stomach, although during an hemoptysis some of the blood may 
be swallowed and this also will be found in the stools. It is important to 
exclude the possibility that the blood in the sputum may have come from 



THE SPUTUM: IN DISEASE 83 

varicosities at the back of the tongue or from lesions in the throat, glottis, 
or esophagus. It is said that the spongy gums of young anemic girls 
explain the blood in their sputum. 

Hemorrhagic Infarction. — Often the diagnosis of a pulmonary infarct 
may be made from the inspection of the sputum alone. The typical sputum 
is expectorated in masses which remain discrete in the cup, some of which 
appear to consist of pure blood, since composed of a very tenacious mucus 
intimately mixed, with fresh blood, while others consist of glairy mucus 
streaked with blood. After an infarction the expectoration begins suddenly 
with cough and pain, or, as was true of half of our cases, there is a sudden 
change in the character of a previous sputum. Microscopically, these 
masses consist of mucus and red blood-cells. Leucocytes are remarkably 
few in number, while alveolar cells filled with blood pigment are usually 
present in enormous numbers. The presence of these epithelial cells, 
however, may be explained by the fact that pulmonary infarctions are 
particularly common in mitral valve diseases. In other cases the sputum 
is much less characteristic, especially if the patient had had considerable 
sputum previous to the embolism. Sometimes, as in one- third of the Johns 
Hopkins cases, there followed a real hemorrhage ; in other cases the sputum 
resembles that of pneumonia, while in still others it is more like the brick- 
red sputum of chronic passive congestion. In one-fifth of these cases the 
thrombosis was followed by practically no sputum at all. (In one, however, 
there may have been some blood-streaked sputum before admission to 
the hospital.) 

The amount of blood in the sputum certainly bears no relation to the 
size of the infarctions. This was well seen in i of our cases with very 
large infarctions and only slightly blood -streaked sputum. All distinctive 
characteristics of the sputum soon disappear, usually in about i week, 
after which time the sputum is merely blood-stained and soon becomes 
more watery and free from blood. 

Chronic Passive Pulmonary Congestion. — In chronic passive pulmonary 
congestion, especially that due to mitral valve disease and particularly to 
mitral stenosis, the sputum is quite characteristic. The patients expector- 
ate, chiefly in the early morning, a sputum which may be uniformly rusty 
in color but which more often consists of a white mucus background in 
which are scattered rusty-colored dots or streaks. This rusty color is due 
to large masses of Hertzfehlerzellen (see page 13). It is in the diagnosis of 
this condition only that the large number and the constant presence of 
these cells have much importance. 

Malignant Disease of the Lungs. — Patients with tumors of the lungs 
and bronchi may have no cough and no sputum but more often it is scanty, 
mucoid or mucopurulent and without distinctive features. An abundant 
purulent expectoration, sometimes putrid, is to be expected after the tumor 
masses have undergone necrosis and cavities develop. Blood is present 



84 • CLINICAL DIAGNOSIS 

eariy in the sputum in from a third to a half of the cases, usually in streaks 
and traces which appear frequently, or it may be so intimately mixed and 
changed that it gives the sputum a rusty color. Frank and even fatal 
hemorrhage may occur. 

The sputum of cases of cancer of the lung has been described as "char- 
acteristically gelatinous, of a red or blackish-red color like currant jelly," 
but this is by no means common and is seen also in non-malignant diseases. 
It more often has a prune-juice character and this Stokes thought an 
important sign. Other patients expectorate a grass-green or an olive-green 
sputum resembling that of caseous pneumonia. In all cases search should 
be made in the sputum for fragments of the tumor. These are more likely 
to be found when the sputum is bloody. Because of secondary infection 
and necrosis these fragments are usually converted into a conglomerate 
mass of degenerated cells and debris without characteristic structure. The 
fresh sputum should be mixed with normal salt solution and examined in a 
flat glass dish against a black background. All masses should be carefully 
teased apart and washed free of adherent blood and mucus. Tumor frag- 
ments may then appear as reddish, grayish or whitish particles or shreds. 

Isolated cancer-cells and cell clusters have been found in the sputum 
from cases with pulmonary carcinoma. Hampeln 64 regards a sputum 
composed exclusively or for the most part of unpigmented polymorphous, 
polygonal cells of variable size, with well-defined nucleus and nucleolus, 
isolated and in clusters, as distinctive of new growth. It certainly is true, 
as he suggests, that the normal squamous cells from the surface of the 
mucous membrane and larynx, also the small polyhedral or cubical cells 
from the deeper layers of the mucosa of the air passages and the polygonal 
alveolar epithelium, are found only in rare instances in the sputum, and 
then only in isolated examples. 

Our cases of this condition presented no unusual points. One case with extensive 
secondary metastasis in the lung had practically no sputum. In another case with a 
large pulmonary tumor the sputum on i day was very viscid, slightly rusty, greenish- 
red in color, not fetid and consisted of pus, red blood-cells and alveolar epithelium with 
much myelin degeneration. Later it was of a dirty grayish color, mucopurulent in char- 
acter and at times contained considerable blood. One case with epithelioma of the 
bronchus had considerable sputum and several severe hemorrhages, while at other 
times his sputum was frothy, seropurulent, liquid and blood-streaked. The diagnosis 
has in several cases been made from tissue fragments found in the sputum. 

Patients with mediastinal growths have cough and sputum if the pres- 
sure of the tumor produces a bronchitis. If the pressure from the tumor 
causes a stenosis of a bronchus, a bronchiectatic cavity will result with 
its profuse fetid expectoration. Gangrene later may develop. 

Syphilis of the Lung. — Syphilis of the lung is a difficult problem to 
discuss with any degree of confidence. Some, especially the rontgenologists, 
find this condition common; others, especially th e pathologists, rare. 

64 Lord, Diseases of the Bronchi, Lungs and Pleura. 



THE SPUTUM: IN DISEASE 85 

How many of the large groups of pulmonary conditions with extensive scar 
tissue formation, often with dilated, bronchi, and which now are predomi- 
nately pyogenic infections, were primarily luetic is difficult to say. Funk 
in McCrae's clinic found 4 cases among 1200 patients who had been thought 
to be unquestionably tuberculous. Fowler and Godley state: ''Evidence 
of excavation with fetid expectoration which does not contain tubercle 
bacilli should always suggest the possibility of the case being one of pul- 
monary lues." The expectoration may be profuse, purulent, and offensive, 
fetor being a common characteristic in advanced cases. These cases usually 
pass as advanced tuberculosis with cavity formation although it should in 
such cases be easy to find Koch's bacillus in the sputum. As a result of 
stenosis of the bronchi, a common event in this disease and due to the exten- 
sive formation of connective tissue at the hilum of the lung, bronchiectatic 
cavities will form and the sputum present all of the characteristics of this 
condition. While hemorrhages are not common some cases attract atten- 
tion by the remarkably bloody nature of the sputum, while some have died 
of hemorrhage. Some writers state that unless repeated examinations for 
the tubercle bacilli be made these cases will pass for consumption . Osier, 
on the other hand, states that he has never seen a case of pulmonary lues 
which clinically resembled tuberculosis. Fortunately in many cases a 
positive therapeutic test will clear up the diagnosis, while already in 1 
case at least Treponema pallidum has been found in the sputum. 

Pneumokoniosis. — According to the dust which is inhaled this con- 
dition has received various names: anthracosis, if it is coal-dust; siderosis, 
if iron-dust; and chalicosis, if it is silicate or other rock-dust. The expec- 
toration is in general mucopurulent, often profuse, and laden with the 
above-mentioned dusts (see page 4). 

Diphtheria. — Cultures should be made from the throat of all patients 
who are suspected to have or recently to have had diphtheria. Cultures 
should certainly be made if any membrane is visible in nose or throat, but 
they should also be made if the throat of one known to have been exposed 
to diphtheria shows a follicular tonsillitis, or even is merely congested. 
If a person known to have been exposed has any constitutional symptoms 
suggesting infection, the bacteriological examination of the throat should 
be made even though it appears perfectly normal. 

Recently a mail carrier, No. 48, admitted July 13, 19 14, came complaining of diffi- 
culty in swallowing and diplopia. He had not been ill, he had not even been feeling 
badly. His throat was merely congested but in the nose and nasopharynx was an old 
slough, in cultures from which Bacillus diphtherise grew. It is of interest that there had 
been an epidemic of diphtheria among the school children along his route. 

Examinations should be made at frequent intervals after an attack of 
diphtheria until at least 2 successive examinations are negative for this 
bacillus. The nasal secretions also of all persons with chronic coryza who 
have been exposed to diphtheria should be examined. Cultures should be 



86 CLINICAL DIAGNOSIS 

made from any membrane forming on a superficial wound in the skin or 
on any mucous membrane. The reason for these careful examinations is 
not nearly so much for the sake of the patient as for the safety of his 
neighbors. In addition to these conditions sometimes diphtheria bacilli 
are unexpected^ encountered. 

G. H., No. 9266, aged 7, was admitted for tonsillectomy. There is no history that 
he had had or had been exposed to diphtheria, but routine examination of the tonsils 
after they had been removed showed the presence of virulent diphtheria bacilli (tested 
by inoculation into a guinea-pig). It is of interest that later one of the nurses who had 
cared for this boy became sick with acute diphtheria. 

Cultures and smears are best made from fragments of the membrane 
(if present) picked from the surface with a pair of forceps. If, however, no 
membrane is seen material for a culture can be obtained on a swab made 
up of a wad of cotton wrapped on the end of a stiff wire about 8 inches in 
length and sterilized in a test-tube. Cultures should not be made from the 
throat within 2 hours after an antiseptic gargle has been. used. 

This sterile cotton swab is forcibly rubbed against the edge of the mem- 
brane if this is visible and, if not, against any exudate or over any injected 
area. Smears are made by rubbing the fragment of membrane or the swab 
on a clear slide, and cultures, by rubbing these forcibly and thoroughly over 
the moist surface of solidified blood-serum. Failures to get a positive cul- 
ture are due frequently to the fact that the swab was rubbed over the 
center of the patch of membrane, and not at the edge, or that the rubbing 
was not forcible enough, or that the surface of the serum was too dry, 
or that the swab was not rubbed firmly enough against the serum. 

The inoculated serum tube is then put into the thermostat as soon as 
possible and left there at 37 C. for from 8 to 20 hours. Smears are then 
made from the growth (for a description of Bacillus diphtherias see 
page 37). 

In addition to Bacillus diphtherias one often finds in the throat Micro- 
coccus aureus and Streptococcus pyogenes. 

The value of the bacteriological examination of the throat is illustrated 
by the fact that McCollum was able to grow this organism from the throats 
of 40% of 500 cases whose throat condition clinically suggested, but was 
not characteristic of, diphtheria, and that in many instances positive 
cultures were obtained from 24 to 48 hours before any membrane appeared. 

Pseudodiphtheria bacilli resemble Bacillus diphtherias, and yet their 
morphology is not quite the same, since they do not show bipolar staining 
and are often shorter and a little thicker than the typical form. They differ 
also in their cultural characteristics and much in their pathogenicity. 
This group certainly includes the pseudodiphtheria bacillus of Hoffmann, 
the xerosis bacillus and others. 

For the bacteriologist this problem is interesting and important, but 
for the clinical laboratorv worker this classification has but academic 



THE SPUTUM: IN DISEASE 87 

interest. He should work only with fresh smears and with cultures less 
than 24 hours old. If using such material he calls Bacillus diphtherias 
all organisms whose morphology is characteristic, he will make fewer mis- 
takes than 1 in 100, and this 1 case will not be hurt if treated for diphtheria, 
while the community will be safer because no chances were taken. 

Vincent's angina, il Plant's angina" "ulceromembranous stomatitis' 1 
and "ulceromembranous angina" are some of the names applied to acute 
or subacute febrile infections of the tonsils or mouth, characterized by the 
formation of deep penetrating ulcers often covered by a pseudomembrane, 
in which large numbers of certain bacteria can be demonstrated. 

To demonstrate these organisms the base of the ulcer is mopped with 
a sterile cotton swab and smears at once made, dried, run through the 
flame 3 times and stained with carbolfuchsin. 

In typical specimens the field will be found crowded with various cocci 
and bacilli and especially with Bacillus fusiformis and a spirocheta. 

It is very important that the student study the flora found in the mouth. 
The number of organisms usually found there is great, over 100 different 
forms having been described. Among these there are some so constant that 
they are considered the natural mouth-flora: bacilli, spirilla, and various 
leptothrix and spirocheta forms, many of them huge, many showing gro- 
tesque involution forms, but all with one common characteristic, that they 
are very hard to cultivate. Among these are Bacillus fusiformis and Spiro- 
cheta dentalis (Miller). Whether any of these are pathogenic or not is 
a question, but one thing is certain, that they increase in great numbers in 
any ulcerative process in the mouth and probably do aid in the formation 
of the ulcers. Undoubtedly they aid in the decomposition of the exudate 
of these ulcers, and explain much of its bad odor. It is very likely that 
Bacillus fusiformis is important in the production of Vincent's angina, but 
the problem is a difficult one, for smears from ill-kept mouths without 
ulcers show such remarkable pictures that the smear from thelbase of an 
ulcer must be very rich indeed in long bacilli and spirochetal before some 
will grant it any importance whatever in diagnosis. 



CHAPTER II 
THE URINE 

GENERAL CHARACTERISTICS 

The Collection and Preservation of Urine. — It is important in all quan- 
titative chemical work which involves the urine that a complete and well- 
mixed 24-hour specimen be collected. In many hospitals the day's collec- 
tion begins at about 6 a.m. The patient voids at this hour and that speci- 
men is discarded. All the urine is then collected until 6 a.m. the next day, 
at which time the patient voids, which specimen completes the collection. 
In case one plans to separate the day urine and the night urine, the former 
period would extend from 6 a.m. to 9 p.m. and the night from 9 p.m. to 
6 a.m., the hours during which the patient is, as a rule, asleep. One then 
estimates the elimination per hour. 

It is necessary that the specimen be collected in a clean bottle and that 
some means be used to prevent the growth of bacteria which under ordi- 
nary conditions is very rapid. If no refrigerator is set aside for this purpose 
a chemical preservative is necessary; which, will depend on the tests to 
be made. If we have chemical work in view we usually use chloroform, 
enough so that at least 1 drop remains undissolved at the bottom. The 
bottle must be tightly corked or bacteria certainly will grow in the upper 
layers of the urine from which the chloroform is volatilizing. This reagent 
adds nothing to the volume of the urine and can be entirely removed. 
Chloroform does not preserve the formed elements, yet it is so satisfactory 
for chemistry that the content of oxybutyric acid in the urine will remain 
unchanged for years. A few crystals of thymol or of gum camphor are 
often used. Formalin is valuable since it preserves the formed sediment of 
the urine. It has, however, two disadvantages; it is an active reducing 
body itself and it adds a sediment of its own. A slight objection to thymol 
is that the urine will give a test suggesting bile. Others add to the urine 
one-fifth its volume of dilute chloroform water or of saturated borax solu- 
tion. The specimens, however preserved, should if possible be kept in an 
ice box. 

Sometimes a 24-hour specimen is not desired. For instance, in the 
diagnosis of slight chronic nephritis a comparison of the urine voided first 
in the morning and of that voided at the end of a day's work gives valuable 
information, or we may ask the patient to exercise violently and examine 
the next voiding. Again, in a mild case of diabetes mellitus only that 
urine voided 3 or 4 hours after a hearty carbohydrate meal may contain 
sugar, and if this voiding were mixed with the entire 24-hour specimen the 



THE URINE: GENERAL CHARACTERISTICS 89 

sugar might be in too dilute solution to be detected. For microscopical 
examination the urine should be studied as early as possible after it is 
voided, and, if possible, without the addition of any preservative. 

The value of the examination of the urine as a routine practice needs 
no emphasis. The perfectly healthy appearance of the patient is no guar- 
antee that the urine will not clear up the diagnosis. The surgeons especially 
need this warning, for all too often i urine examination would probably 
have prevented an operation following which the patients have died in 
diabetic coma. 

The Amount of Urine. — The limits of the daily amount of urine to be 
considered normal vary widely. Generally speaking we say that an adult 
should not void less than 800 c.c. or more than 3000 c.c. per 24 hours. 
The average output is usually stated as from 1 500 to 2000 c.c. That may be 
true for a country in which beer drinking is common, but in this country 
from 900 to 1500 are given as normal limits. The average daily output 
of women is slightly less than of men. The amount of urine depends in 
part on the size of the person. That of an adult is almost directly propor- 
tional to his weight, but children excrete relatively more than do adults; 
newly born infants void from 150 to 200 c.c. a day, and children from 3 to 
5 years of age about 700 c.c. 

The amount of urine per day in a normal person depends chiefly on the 
volume of fluids ingested. By varying his fluid intake this may vary from 
800 to 3000 c.c. per 24 hours. The increase in the output after drinking 
a large amount of water at one time reaches its maximum in from 2 to 3 
hours and lasts from 5 to 6 hours. Yet, as all have experienced, the water 
output is as capricious as is that of the other urinary constituents. 

The margin of functional ability of the kidney is surprisingly large, 
as is seen in diabetes mellitus, in which disease a practically normal kidney 
may eliminate each day 2 5 liters of water, an absolutely increased amount 
of the normal solids and several hundred grammes of sugar and will endure 
this increased work for a long time without showing any sign of disease. 
In the case of a rabbit Kulz was able by intravenous injection of salt 
solution to increase the output of urine to 256 c.c. per hour for 9 hours, 
and yet the qualitative composition of this urine remained normal. Insen- 
sible as well as copious perspiration affects the amount of urine, and so, 
other things being equal, it will be greater in cool weather than in hot. The 
amount voided in health as well as in disease is also affected by the loss of 
fluid in other ways, particularly by diarrhea and by vomiting. 

The water content of exudates (pleural or ascitic), of subcutaneous 
edema and of other abnormal accumulations in the body is finally excreted 
through the urine. This explains the polyuria seen in nephritis while the 
edema is disappearing. To demonstrate this the person should be put on 
constant diet and constant fluid intake and the urine carefully measured 
or the increased output may pass unnoticed. 



90 CLINICAL DIAGNOSIS 

The differences in the hourly amounts of urine voided during the day 
and during, the night has not received the attention it deserves. Quincke 
and his students found that in liver, kidney, and in heart diseases which 
produce edema the urine voided per hour during the night is greater in 
amount and contained more solids than that during the day, a condition 
sometimes called nycturia. Normally the reverse is true; the kidneys seem 
to sleep with the rest of the body and secrete an amount per hour during 
the day which is to that per hour during sleep as ioo : 50 to 60 or perhaps 
as 100 : 80 to 90. In cases of cardiac or arterial disease and in nephritis 
the reverse is true. This is called " the fixation of specific gravity." It 
would seem in such cases as though the kidney improved its opportunity 
during the sleeping hours to eliminate that which it could not during the 
day. In a well-marked case of nephritis, D : N : : 100 : 200 and in 1 case x 
which we followed the ratio was even 100 : 544. This does not depend, 
we are convinced, on changes in the position of the patient's body and 
therefore on changes in circulation depending on this. This test has definite 
diagnostic value since it aids to differentiate cases of functional circulatory 
and renal disturbance (e.g., hysterical) from organic diseases. The dis- 
turbance of this ratio is particularly marked in case the output of urine 
is increased, as in diabetes, or by diuretics or by exercise during the day. 
The disturbed ratio is not present in heart disease providing the compen- 
sation is good. Cardiac insufficiency seems the underlying cause in most 
cases of the disturbed ratio. 2 

By polyuria is meant an output of over 3000 c.c. of urine in 24 hours. 
An output of 800 c.c. or less in 24 hours is termed an oliguria. To be 
important clinically an increase or diminution in the output of urine should 
extend over several consecutive days. These limits are quite elastic, even 
for normal persons. 

Pathological Factors Influencing the Amount of Urine. — (1) 
The condition of the renal parenchyma in diseases with bilateral diffuse 
lesions. A general rule is that the more acute the nephritis the less the 
amount of urine excreted. In acute nephritis there may at first be anuria, 
or 50 to 100 c.c. only per day; in a subacute nephritis about a normal 
amount, while in a chronic interstitial nephritis the patient may void from 
6 to even 12 liters each 24 hours. 

(2) The velocity of the blood-current through the kidney determines 
in great degree the amount of urine, the general law being that the amount 
of urine varies directly as the amount of blood which passes through the 
kidney in a unit of time; that is, as the rapidity of the blood-flow, and not 
as the blood-pressure. Hence chronic passive congestion of the renal 
circulation, whatever -the cause, diminishes the output and drugs which 
improve the renal circulation increase the output and so act as " diuretics." 

1 Johns Hopkins Hosp. Rep., vol. x, p. 323. 

2 Lasoeyres, Deut. Arch. f. klin. Med., August 16, 1900. 



THE URINE: GENERAL CHARACTERISTICS 91 

(3) Disturbed Metabolism. — The output of urine depends much on the 
quality and the quantity of the substances to be eliminated. The best 
illustration of this is the patient with diabetes mellitus who may void 
even 2-5 liters of urine per day when his sugar output is high and when, by 
modifying his diet, the sugar output is reduced, the water output, dimin- 
ishes as well. The so-called " epicritical polyitria " which occurs after 
fevers may have a similar explanation although we do not know the sub- 
stances involved. Some cases of typhoid fever, for example, during con- 
valescence void from 4 to 6 liters of urine per day, and the same may be 
true of almost any disease which has diminished the output of urine. This 
is beautifully seen in cases of subacute parenchymatous nephritis. In 
such cases a polyuria accompanies the elimination of substances which 
were retained during the acute period of the illness and indicates a 
favorable prognosis. 

(4) Psychical disturbances and various nervous storms may be followed 
by polyuria, as may also angina pectoris, hysteria, and epileptic convul- 
sions. The cause is probably vasomotor. The so-called " paroxysmal 
polyuria ' ' is probably a functional disturbance. 

(5) Another cause of periodic polyuria is the hydronephrosis seen in 
movable kidney, etc. 

(6) Chilling of the skin may be followed by polyuria. 

(7) Of the various forms of polyuria the causes of which are unknown, 
diabetes insipidus is the most striking example. A patient with this disease 
may void even 1 2 or more liters in a day. 

(8) Hydremia. 

(9) Lesions of medulla. 

(10) Stimulation of renal secretion by drugs or " renal diuresis " due 
to the action oi e.g., the caffein group. 

It is often of interest to estimate the proportion of water intake which the urine 
represents. The normal person eliminates from 60 to 70% of water intake through the 
kidneys. If he increases greatly the amount of water consumed the bulk of the increase 
will appear in the urine and the percentage of urine relative to total intake may rise 
to even 96%. In 2 cases of chronic interstitial nephritis however the output of water 
through the kidneys was relatively high even though absolutely small. One of these 
cases with an intake of i960 c.c. voided 85% and on the next day, of the 2400 c.c. he 
drank, he eliminated 86% through the kidneys. In another case, of 1370 c.c. of fluid 
ingested, 85% was voided in the urine and on another day 83% of 1790 c.c. In chronic 
parenchymatous nephritis, with the patient in almost stable condition and receiving 
exactly the same amount of fluid each day for 26 days (6200 c.c), the average daily 
output by the kidneys was 66%. In cases with ascites and signs of renal insufficiency 
the renal output will drop to 40% of the intake and in anuria even to o. The follow- 
ing figures from a recent case of eclampsia in the obstetrical ward illustrates this 
well. The patient was not urged to drink much. On the first day after the con- 
vulsions, of 8350 c.c. of water drunk the kidneys excreted 20%; the next day, of 
IO >535 c.c, 80%; on the fourth day of 9400 c.c, 93%; and on the fifth day the patient 
drank 7100 c.c. of water and voided 7390 c.c. of urine. During this time there was 
some diarrhea. 



92 CLINICAL DIAGNOSIS 

Anuria, or the absence of micturition, whether from failure of renal 
secretion or from retention of the urine, may be due to a variety of causes, 
which may be classified as obstructive, paralytic, septic, renal, and prerenal. 

The best illustration of the obstructive type of anuria is that seen in 
tumor or hypertrophy of the prostate gland, in vesical calculus, or in trauma 
of the urethra. The paralytic form follows transverse lesions of the spinal 
cord, but the case which develops early and insidiously in typhoid fever 
deserves particular mention since it is unfortunately so often overlooked. 
The reflex type is best illustrated by cases with a calculus in the pelvis 
or ureter of i kidney, or an operation, e.g., nephrectomy of i kidney and 
reflex inhibition of the secretion of the other. 

The most important form of anuria is that due to renal disease. In 
all cases of acute nephritis there is a reduction of the urinary output which 
varies in direct proportion to the intensity of the acute process. In such 
cases usage has permitted the term anuria to include cases who void a 
little but negligible quantity (e.g., ioo c.c. or less) in 24 hours. 

The prerenal causes are numerous. This form of anuria may be due 
to purely functional causes, as hysteria, in which case it is temporary and 
is followed by a polyuria; or to certain fevers, as scarlet fever; to cert air. 
poisons, x as phosphorus, lead, turpentine, ether and chloroform; it occurs 
in collapse; and often, but not always, with approaching death. 3 The 
anuria of Asiatic cholera is attributed to inspissation of the blood. It is 
surprising how long a person can live after all renal tissue has been removed 
or destroyed or when there is complete suppression of renal function. 
Moxon reported a case of ureteral calculus with anuria lasting 14 days, 
which recovered after the passage of the stone. Adams' patient had anuria 
for 19 days and yet recovered. Polk's patient lived for 11 days after his 
1 and only kidney had been removed. 

Specific Gravity. — The specific gravity of the urine is its weight com- 
pared with that of an equal volume of water. The standard is the weight 
of a liter of water, 1000 gms. (some say, of a cubic centimeter of water, 
1.0 gms.). If urine has a specific gravity of 1018 (or 1.018) we mean that 
a liter of it would weigh 1018 gms. (and 1 c.c, 1.018 gms.). The specific 
gravity of any fluid may be determined accurately by weighing a given 
amount of it in a pycnometer, but clinically it is determined by measuring 
its buoyancy by means of a form of aerometer called a urometer. The 
aerometer spindles are usually graduated from 1000 to 1050. It is better 
to use 2, 1 graduated from 1000 to 1020, the other from 1020 to 1040. The 
practitioner should be careful to get good instruments, for some on the 
market are very inaccurate, especially those of the smaller types. The 
glass cylinder to be used should have parallel sides, fluted if possible, a 
wide base and a good spout. This is filled about % full of urine and the 
foam, if present, removed with a piece of filter paper. The bobbin is then 

3 See also Bevan, Am. Surg., April, 1903. 



THE URINE: GENERAL CHARACTERISTICS 93 

dropped in and allowed to come to rest. The observer should now assure 
himself that it actually floats and does not touch the side of the cylinder. 

While making a reading the eye should be on the level of the base of the 
meniscus. Two or 3 separate readings should be made, the bobbin being 
pushed down each time and then allowed to come to rest. It is very import- 
ant to make the proper corrections for temperature. These instruments are 
standardized usually for 1 5 C. A difference in temperature of 3 C. means 
a difference of 1 in the fourth place of the specific gravity reading. That 
is, a urine which at 15 C. has a specific gravity of 1.012, will at 18 C. 
read 1.011. This correction is usually of slight importance if the urines 
are of average concentration, yet we suspect that failures to consider it 
explain the impossibly low figures of the specific gravity of the urine of 
certain cases of diabetes insipidus and chronic interstitial nephritis. This 
correction must of course be carefully determined if the specific gravity 
is to be used as the basis of quantitative work, as for instance, the estima- 
tion of the total solids or the amount of sugar or of albumin. It is only 
§&U to say that for the latter we think the aerometrical methods at their 
best are hardly accurate enough, and that the urine should be weighed on 
a good chemical balance. Again, an instrument suited for salt solutions 
is not always accurate in a sugar or albumin solution. 

In recording the specific gravity of urine mention should always be 
made of the character of the specimen examined, when voided, etc. In 
general, only a well mixed 24-hour specimen should be tested, for the 
specific gravity of the various voidings during the day and night may vary 
from 1.002 to 1.040, depending on the food, the fluid, the lungs, the skin, 
etc. It may be very high after severe exercise with sweating, after transu- 
date formation, etc. Two normal men recently were refused on first 
examination by life insurance companies because they happened to have 
eaten food just before examination which for them had a diuretic action 
and so the specific gravity of the urine was abnormally low, in 1 case 1.003. 

The specific gravity of the mixed 24-hour urine of the normal adult 
varies from 1.015 to 1.020. In the new-born from 1.005 to 1.007. 

In case the amount of urine is too small to fill the cylinder, it may be 
diluted to a known volume. The formula for the correction is: Sp. gr. = 
1.000+ab, in which " b"=the dilution, and " a " the last 2 figures of the 
specific gravity found. For instance, if the urine was diluted with just 
twice its volume of water and if the reading of the diluted urine was 1.006, 
Sp. gr. = 1.000+3X6 = 1.018. 

In some cases it is the specific gravity of the single voidings which are 
desirable. For instance, in the diagnosis of an early chronic diffuse nephritis 
the constantly low specific gravity of the morning urine is of value (see 
page 312). 

The specific gravity of the urine depends chiefly on the relative amounts 
of water, urea and sodium chloride which it contains. The amount of 



94 CLINICAL DIAGNOSIS 

water will depend on the factors discussed on page 90. A high percentage 
of urea explains in large measure the high specific gravity of the urine in 
fevers. The amount of salts is increased by foods, by the medicines taken 
and by the absorption of transudates. While in general the specific gravity 
of the urines of any 1 person will vary inversely as its amount, this is not 
strictly true since an increased output of water increases the output of 
solids. A marked exception to the rule is diabetes mellitus, in which disease 
the urine is greatly increased in amount and also in specific gravity ; while 
another exception and in the opposite direction is seen in nephritis with 
renal insufficiency, in which case there is oliguria and a greater diminution 
in the output of solids than of water and therefore a low specific gravity. 
In nephritis a definitely low specific gravity is rather suggestive of an 
impending uremia. It is also seen in cases of malnutrition in which the 
metabolic processes are at low ebb, as for instance, in a patient of Chabrie, 
a girl of 20 years of age, whose output of urine on 1 day was but 750 c.c. 
with a specific gravity of 1.008. In diabetes insipidus the specific gravity 
is very low (some ridiculous figures are probably explained by the failure 
to make a correction for temperature) . 

An approximate estimation of the total amount of solids in the urine 
may be made by the use of Haser's coefficient, 2.33. The last 2 figures of 
the specific gravity multiplied by this empirical coefficient will give fairly 
accurately the number of grammes per liter of solids excreted. 

Authorities disagree concerning this coefficient. Neubauer gives 2.328. Donze 4 
states that the coefficient should be slightly lower for dilute than for more concentrated 
urines and therefore should vary from 1.850 to 2.440, with an average of 2.210. 

The specific gravity of the urine may be used in the quantitative esti- 
mation of sugar, albumin, etc. If used for this purpose, however, a good 
urometer should be used and the temperature correction carefully made, 
otherwise the result may be absurd. 

Color. — The color of the urine of a normal adult is usually a shade of 
yellow, its depth varying with the dilution of the urine and hence directly 
with its specific gravity, a dilute urine being of pale and a scanty urine of 
dark yellow color. Exceptions to this are : diabetes mellitus, in which case 
the urine is very pale and yet is increased in amount and has a high specific 
gravity, a point which will sometimes suggest the diagnosis; in aplastic 
anemias, especially chlorosis, in which cases the urine is pale from lack of 
pigment, while in those anemias in which there is increased destruction 
of the red corpuscles, as in pernicious anemia, the urine is highly colored. 
As a rule acid urine is more highly colored than is alkaline urine. In 
uremia the urine often is very pale, a fact which was responsible for the 
old theoiy that a retained pigment is the cause of this condition. In cases 
of certain grave infections which seem to have destroyed the bile-producing 

4 Compt. rend. Soc. de Biol., 1903, 155, 537. 



THE URINE: GENERAL CHARACTERISTICS 95 

function of the liver the urine is said to have been without any pigment at 
all. A febrile urine is dark since it is concentrated and also because it 
contains more of uroerythrin and other pigments than does normal urine. 
Urine turns dark on being exposed to sunlight. For this reason the con- 
trast in color between the day and night urine is often striking, the day 
urine being of a golden-yellow and the night of a pale green color. This 
difference is due partly to the amount of pigment excreted but more to 
the effect of sunlight which changes the colorless chromogens to pigments 
in the specimen collected during the day. 

The use of a color scale is recommended in order to avoid the variety 
of terms used to describe the same color (yellow, light yellow, amber, 
orange, straw, etc.). 

The pigments normally present in the urine are : 

Urochrome, the predominant one, gives urine its normal shade of 
yellow, orange or brown, according to the amount present. It has not 
yet been isolated and so its empirical formula is not yet known. Indeed 
several pigments may be included under this name. It has no absorption 
spectrum and no fluorescence. There is evidence that it is derived 
from urobilin. 

Hematoporphyrin in small amounts is present in every normal urine 
(see pages 97 and 242). 

Uroerythrin is often present under normal conditions and explains the 
salmon-red color of the urate sediment. It is increased in amount by a 
rich meat diet, by profuse sweating, alcoholic drinks, violent exercise and 
by certain digestive disturbances. It is increased also in fevers, in circula- 
tory disturbances and in arthritis. This pigment may be demonstrated by 
shaking the urine out gently with amyl alcohol, which will become orange 
in color and will give the characteristic spectrum of uroerythrin. This 
pigment bleaches in a characteristic manner on exposure to light. When 
dissolved in concentrated sulphuric acid its solutions are carmine-red, which 
on the addition of an alkali changes to purple, then to blue and finally 
to green. 

Urobilin is a constituent of the normal urine, in amounts varying from 
30 to 120 mg. per day. Urobilin itself is not present in perfectly fresh 
urine, but its chromogen, urobilinogen, is, and this on exposure to sunlight 
yields urobilin. In the following lines we shall include under the term 
" urobilin " this pigment and its chromogen or chromogens. 

Whether urobilin is a single pigment or a group of pigments is a doubtful 
question. It has been impossible to isolate it without some decomposition 
and all efforts to remove its impurities have thus far failed. 

The origin of the urobilin present in normal urine is still in doubt, 
but evidence favors the theory that it originates in the intestines and in 
the liver. There is no constant relation between urobilinuria and urobil- 
inemia, or between bilirubinemia and urobilinuria. The attractive theory 



96 CLINICAL DIAGNOSIS 

of Gilbert and Herscher and others 5 that the kidneys transform the bili- 
rubin of the blood into the more diffusible urobilin has not received con- 
firmation. Conner and Roper 6 found that bilirubinemia and urobilin uria 
bear a rough quantitative relation to each other and yet their work furn- 
ishes little support to the theory that the urobilin of the urine originates 
in the kidneys. Certainly a great deal of urobilin is formed in the intestine 
(enterogenous formation) as the result of the reducing action of certain 
bacteria on bile pigment. It seems to be identical with stercobilin. 

Some urobilin is said to be formed in areas of hemorrhage into the 
body tissues (histogenous formation), and, indeed, wherever there is 
blood destruction from any cause (hematogenous formation) as after 
toxic doses of blood-poisons, such as antifebrin and antipyrin. Meinel 7 
found that a certain amount is formed in the stomach in some cases 
of hyperacidity. 

The urobilin of the urine is increased also in fevers, in chronic passive 
congestion, lead poisoning, atrophic cirrhosis of the liver, etc. It is increased 
before and after a period of obstructive jaundice. There is a life-long 
increase in persons with chronic family jaundice (Tileston and Griffin). 

When there is a marked urobilinemia there may be also a definite 
urobilin jaundice. 

Urobilin does not give Gmelin's test; it gives a test similar to the biuret ; 
and if to a urine made strongly alkaline with ammonia and filtered be added 
a i% alcoholic solution of zinc chloride the presence of urobilin will be 
indicated by a beautiful green fluorescence and spectroscopically by the 
characteristic spectrum of alkaline urobilin. The spectrum of acid urobilin 
may be obtained with urine to which have been added a few drops of a 
mineral acid, but it is better to shake it out with amyl alcohol and then 
examine the extract, or to add an equal amount of 10% ZnAc in absolute 
alcohol, and filter. 8 This test will be given even in spite of the presence of 
considerable bilirubin. The fluorescence is best seen with a convex lens 
which gives a luminous green circle. 

For the quantitative determination of urobilin Hoppe-Seyler recommended to acid- 
ulate ioo c.c. of urine with sulphuric acid, saturate it with ammonium sulphate and 
allow it to stand for some time; then filter and wash the precipitate with saturated 
ammonium sulphate solution. The precipitate is then pressed out between blotting 
papers and extracted repeatedly with equal parts of alcohol and chloroform. The 
extract is then filtered into a separating funnel and to the nitrate are added 2 volumes 
of water and then chloroform until the chloroform settles out in a clear layer. The 
chloroform solution is evaporated on a water-bath and the residue dried at ioo° C. 
It is then extracted with ether, the ether extract filtered off, the residue dissolved 
on the paper in alcohol, again brought into the weighed beaker, evaporated, dried 
and weighed. 

5 Compt. rend. Soc. de Biol., 54, p. 795. 

6 Arch. Int. Med., Jan., 1909, vol. ii, p. 532. 

7 Centralbl. f. inn. Med., 1903, vol. xxiv, p. 321. 

8 Schlesinger, Deutsch. med. Wochenschr., 1903, No. 32, p. 561. 



THE URINE: GENERAL CHARACTERISTICS 97 

Among other chromogens in the normal urine are indoxyl-sulphuric 
acid (see page 143), indoxyl-glycuronic acids and possibly skatoxyl-sul- 
phuric and skatoxyl-glycuronic acids. Among the pigments which may 
under pathological conditions be present are hemoglobin, methemoglobin, 
hematin, bile pigments, melanin and others; some come from drugs, e.g., 
chrysophanic acid; others from the foods, e.g., the pigments of various 
berries, cherries, etc. 

Blood. — The color of a urine which contains blood will depend on the 
amount of blood present and on the modifications which the blood-pigment 
has undergone: hemoglobin gives the urine a reddish tint, methemoglobin 
a brownish one. When only a little blood is present the urine often has a 
characteristic smoky tint due to methemoglobin. When more is present 
its color may be reddish-brown, brown, almost black, or greenish-black, 
as in the black-water fever of hemoglobinuria. Such urine is cloudy because 
of the large number of blood corpuscles and other organized elements of 
sediment usually present. If the sediment is very heavy one may find in 
it masses of amorphous hemoglobin. 

Hematoporphyrin. — Hematoporphyrin may be present in the urine 
in large amounts after the long continued use of trional, sulphonal, or 
tetronal; also in cases of typhoid fever. Thick layers of such urine have 
a dark or blackish color; thin layers, a yellowish-red or violet color. The 
black color sometimes seen in such cases is, Garrod thinks, due only in 
part to hematoporphyrin and more to some unstable purple pigment. 

Bile. — The urine of the jaundiced patient usually contains bile pig- 
ment, but in cases of very mild jaundice urobilin alone may be present. 
The color of the urine which contains bilirubin and biliverdin may be dark 
yellow, brown, green, greenish-black, or in long-standing cases even quite 
black, depending on the amount of bilirubin which has become changed to 
biliverdin and other modifications of this pigment. If an acid urine which 
contains considerable bilirubin be allowed to stand in a cold room a sedi- 
ment of bilirubin in needle crystals may be deposited. It is often possible 
to detect the presence of even very small amounts of bile in the urine by 
shaking this enough to produce a foam. This foam, white in all other 
urines no matter how dark they may be, is stained yellow by bile ; it may 
also be yellow if very much urobilin is present. 

Melanin is present in the urine in cases of melanotic tumors which 
have invaded the viscera, especially the liver (Garrod). Such urine may 
be black but more often is quite normal in color when voided, since the 
pigment is then present as the colorless chromogen, which later is split 
by sunlight, yielding melanin. The change in color begins at the top and 
extends downward through the urine forming sometimes a sharply defined 
black layer above one of colorless urine. This transformation of melanogen 
may be hastened by the addition to the urine of nitric acid or of any other 
oxidizing body, especially ferric chloride, which will at once turn the urine 
7 



98 CLINICAL DIAGNOSIS 

black and throw down a gray precipitate which is soluble in excess of this 
reagent. This, the ferric chloride test, is the most delicate and reliable 
of all tests of melanin and necessary for its recognition. 

Homogentisinic acid, the chief pigment present in the condition 
named " alkaptonuria," gives to the clear urine after standing or 
after the addition of an alkali (see page 208) a brownish-black color and 
a syrupy consistency. 

The urine is sometimes very dark, almost black, in cases of peritonitis, 
gangrene of any organ, and in any condition, including simple constipation, 
which favor the formation of the aromatic products of decomposition, the 
ethereal sulphates of indoxyl, etc. In these cases the urine is sometimes 
very blue, not from indigo but from higher oxidation products of indol. 
Such urines, clear at first, will blacken if nitric acid be added and they 
are then warmed. They do not blacken on the addition of ferric chloride 
and do not reduce copper solutions. 

Some urines are very dark when voiding, others only when they have 
stood a long time. This color may be due to pyrocatechin, C 6 H 4 (OH) 2(1.2), 
which in alkaline solution is oxidized by the air to a greenish-brown and 
finally black color. Such urine will reduce alkaline copper sulphate if 
heated but will not bismuth. Some {e.g., Baumann) believe that pyro- 
catechin is derived from the vegetables of the food. 

To isolate pyrocatechin the urine is concentrated by heat, then filtered, 
a little sulphuric acid added, and then boiled to drive off the phenol. 
It is then shaken out repeatedly with ether; the ether is distilled off, the 
residue neutralized with barium carbonate and again shaken out with 
ether. The ether is then allowed to evaporate and the pyrocatechin will 
crystallize out. 

Hydrochinon, C 6 H 4 (OH) 2(1.4), is present in the urine after the use 
of phenol. Its decomposition products give to the urine a dark color and 
reduce copper solutions easily. 

Urine containing indoxyl in large amounts is clear when voided, but 
soon becomes dark from the presence of indigo. The blue of the indigo 
may be masked by the yellow color of the urine. The scum of such a urine 
may be blue. Sahli mentions such a case, that of a boy whose urine when 
voided was of a green-grass color. 

Ochronosis is a rare disease characterized by blackening of the cartilages. 
The urines of these patients turn black on standing. Osier reported 2 
such cases whose urine contained the alkapton bodies, but it is said that 
in other cases the black color of the urine is due to other pigments. 

Garrod 9 classifies the causes of black and very dark urines as follows: 
long-standing jaundice; hematuria or hemoglobinuria; melanotic sarcoma; 
alkaptonuria; ochronosis; indoxylsulphate in great abundance; certain 
cases of tuberculosis in which cases the urine must stand for some time, 

9 The Practitioner, 1904, vol. lxxii, p. 383. 



THE URINE: GENERAL CHARACTERISTICS 99 

even a month before the color develops; certain drugs, as phenol; and rare 
cases due to unknown pigments. Of these cases in only 2 is there really 
black urine : melaturia, and alkaptonuria on standing. 

In chyluria the urine has a milky appearance. 

Colors Due to Medicines. — The list of medicines which may modify 
the color of the urine is too long to tabulate. In general it may be said 
that if the urine of a patient has an unusual color inquiry should always 
be made concerning the previous medication. Among the drugs which 
deserve mention are carbolic acid, whether applied internally or externally 
(in which case the color of the urine is important to control therapy), tar 
preparations, resorcin, naphthol, salol, and many aromatic bodies. The 
change of color may appear only if the urine is alkaline and has stood for 
a long time. Methylene blue even in small amounts, e.g., 0,1 gm., will 
color the urine for several days. In 1 hour after the dose the urine has a 
greenish color, later a deeper green, then a blue, which may last even 3 or 4 
days. This color may be intermittently present, e.g., only in the first morn- 
ing voiding. It may be intensified or even be made apparent by boiling the 
urine after adding acetic acid since the pigment is in part voided in colorless 
form. Weber 10 thinks methylene blue explains practically all the blue and 
green urines and doubts that any are due to indigo blue. He emphasizes 
the common use of methylene blue to color candies and food-stuffs. 

Drugs containing chrysophanic acid, e.g., chrysarobin, rhubarb, san- 
tonin, senna, and others, give the urine a yellow tint when acid and a red 
tint when alkaline. The pigments of many vegetables and fruits will 
change the color of urine, e.g., turnips, whortleberries, blackberries, etc. 

Odor. — The odor of normal fresh urine is not unpleasant. The unpleas- 
ant so-called " urinary " odor is due to the ammoniacal decomposition of 
urea by bacteria. That of a decomposing albuminous urine may be espe- 
cially disagreeable. The urine of patients with cancer of the bladder and 
deep inflammatoty disease of the urinary tract may have an intolerable 
odor. Chabrie believes that the urine has a characteristic odor in certain 
abnormalities of metabolism, such as diabetes and oxalura. We may even 
imagine from his writings that he thinks that one of the great masters of 
French medicine could diagnose insanity from the odor of the urine alone. 
The urine is said to have a special odor in chyluria and even in hematuria. 
Sometimes the urine has a remarkable absence of odor. We have noticed 
a strong odor of H 2 S in the quite fresh urine of certain nephritics. It 
should always be remembered, however, that the bottle in which the 
patient brings the specimen may explain the odor. 

Certain odorous substances are excreted as such in the urine, e.g., 
valerian, asafetida, coffee, and various foods. Others build odorous bodies, 
e.g., balsams, copaiba, cubebs, etc. Turpentine gives the urine the odor 
of violets; asparagus that of methyl-mercaptan. 

10 Lancet, September 21, 1901. 



100 CLINICAL DIAGNOSIS 

General Appearance. — The fresh urine of a normal person is quite clear. 
The one exception is the so-called phosphaturia (see page 101). A faint 
cloud, named the nubecula, soon appears in the upper layers of a clear 
urine, which consists of mucous strands enclosing a few cells. After stand- 
ing, any normal urine may become cloudy; if acid, from a urate sediment 
(see page 245), if alkaline because of the rapid growth of bacteria which 
form ammonia from the urea, from a precipitate of the phosphates. 
In pathological urines a cloud when the urine is perfectly fresh may 
be due to bacteria, to precipitated phosphates or to an abundant 
organized sediment. 

Reaction. — The fresh urine of a normal person is acid or amphoteric; 
in certain cases of phosphaturia it is alkaline. The quantitative determina- 
tion of the reaction of the urine has proved a very attractive field but the 
results are far from satisfactory. Until recently the " degree of acidity " 
of a solution was understood to mean the amount of hydrogen which could 
be replaced by the metal of an alkaline solution (NaOH) regardless of the 
previous state of the hydrogen, whether free as hydrogen ions or in combina- 
tions which could be easily disassociated and the hydrogen substituted for 
by the metal of the alkali. 

The physical chemists define " degree of acidity " as the absolute 
number of disassociated H-ions in each liter of urine. Judged by this 
standard urine is only about 30 times as acid as is distilled water and only 
about Xo as acid as titration would indicate. Is the determination of 
either of these " degrees of acidity " of value and if so, which? One great 
difficulty is that that acidity determined by titration is due to a consider- 
able number of chemical substances, the most of them acid salts, and hence 
the question of color indicator is a very serious one, since the points indi- 
cated by the various ones as the neutral point differ much. Phenolphtha- 
lein is the indicator usually used. This has as practical advantage the 
sharpness of its end reaction and the fact that of the indicators it itself 
is the weakest acid. But it is a poor indicator, perhaps the worst, in the 
presence of ammonium salts. Whatever results are obtained with it 
have not an absolute but an empirical value. In the case of man the 
reaction of the well preserved and well mixed 24-hour specimen of urine 
is always faintly acid to litmus to a degree corresponding to about 1.15 
to 2.3 gms. of HC1 for each 24 hours. This acidity depends chiefly upon 
the diet and is greater the more the proteid ingested. The urine of herbiv- 
orous animals is alkaline since the organic acids of their food are oxidized 
to alkaline carbonates. Yet if these animals are starved their urine will 
be acid to litmus, since their tissue proteid then becomes their chief food. 
The urine of a man on a vegetable diet will be less acid, or even amphoteric, 
from the increased ingestion of alkali-forming foods. 

In no case is there any free acid in normal urine, but rather acid salts, 
especially diacid sodium phosphate, and many others produced by the oxi- 



THE URINE: GENERAL CHARACTERISTICS 101 

dation of neutral proteids. Among these are salts of sulphuric, phosphoric, 
hippuric, oxalic, and the oxy aromatic acids. Just what part each of these 
plays in the acidity of the urine, however, cannot be decided. Certainly 
uric acid is not a factor, since its solution is neutral to litmus. 

The urine of a starving man may have an acidity of constant value, 
but that of others shows constant variations due especially to the diet 
and known as the " alkaline tide." The acidity is highest in the morning 
before breakfast and lower for a few hours after each meal, especially after 
the breakfast, due to the secretion of the hydrochloric acid of the gastric 
juice. The acidity is later restored to its previous value when the hydro- 
chloric acid is reabsorbed. 

For a short time after a meal, from 2 to 4 hours, the urine may even be 
alkaline when freshly voided and hence turbid with a sediment of phos- 
phates of the alkaline earths. 

Phosphaturia is the name given to an interesting symptom-complex 
characterized by the presence of a heavy precipitate of the earthy phos- 
phates in the freshly voided urine. Formerly, as the name would imply, 
this was supposed to be due to an increased output of phosphoric acid. 
There is, however, no such increase and the precipitation is due rather to 
a change of reaction, for the phosphates can remain in solution only in acid 
medium, so that the name " alkalinuria " would be much more suitable. 
Phosphaturia {i.e., alkalinuria) may be present if the diet consists of 
vegetables ; in cases of gastric diseases with considerable loss of hydrochloric 
acid to the body through vomiting or lavage and perhaps through diar- 
rhea also; and in a group of nervous patients without either of the above 
mentioned causes. In this last mentioned group the phosphoric acid out- 
put during the periods of " neurasthenia " with phosphaturia has been 
found diminished to about half its normal value, the nitrogen output 
decreased, but the calcium output increased. The trouble would seem to 
be an excess of the output of calcium relative to that of phosphoric acid. 
In Soetbeer and Krieger's case the phosphoric acid output was practically 
normal, the calcium was increased even to 0.7 gm. a day (normal 0.2) and 
Ca : P2O5 : : 1 : 1.5 to (normally 1 : 12). Some cases 11 would seem to 
have during their periods of phosphaturia symptoms definitely referable 
to or coincident with this abnormal metabolism, but due to changes in 
calcium metabolism rather than to that of the phosphoric acid. In 1 case 
the calcium output was increased more than 3-fold, perhaps as a result 
of chronic colitis. Phosphaturia occurs also after sexual excesses and 
during the periods of depression following psychical exaltation. Freuden- 
berg 12 carried this idea to an extreme, separating phosphaturia, latent 
phosphaturia (in which the phosphate precipitate appears when the fresh 
urine is heated) , and ammonuria (in which case moist litmus held over the 

11 Soetbeer and Krieger, Deutch. Arch. f. klin. Med., 1902, vol. lxii, p. 553. 

12 Deutsch. med. Wochenschr., September 17, 1903. 



102 CLINICAL DIAGNOSIS 

mouth of a tube of heated urine will turn blue). He thinks that these are 
3 grades of the same abnormality, which is found in sexual neurasthenics 
especially but not in patients with true hysteria. It is often met with among 
mental cases (Heinicke). 

There are a few cases with general neurasthenic symptoms in whose 
urine the phosphoric acid is definitely increased, and who later perhaps 
will develop polyuria or glycosuria. Senator suggests that some of the 
cases of diabetes insipidus, the specific gravity of whose urine is higher 
than in other cases, may belong here. 

The reaction of the urine can be much modified, even made alkaline, 
by drugs, particularly by large doses of the alkaline salts. Milk of lime 
in sufficient doses will make the urine alkaline from the presence of ammon- 
ium carbamate (Abel). While a transudate is being absorbed the urine 
may be alkaline, also after a hemorrhage into the intestine, in which case 
it is due to the absorption of the blood-salts. The urine is alkaline also 
in certain cases of pneumonia, typhoid fever and in diseases of the central 
nervous system. We have noted a marked alkalinity in certain cases of 
nephritis, particularly of the severe chronic parenchymatous type with 
much edema, which renders the examination for casts difficult. 

The urine may be alkaline because of the alkaline secretions and exu- 
dates of cystitis or urethritis and, lastly, because of alkaline fermentation 
due to the action of bacteria in the bladder which break up the urea into 
ammonium carbamate and carbonate. To determine whether the alka- 
linity of the urine is due to a fixed alkali or to ammonia (in which case, it 
always is the result of bacterial fermentation) a strip of red litmus paper 
is wet with the urine and then dried. If the red color returns the alkalinity 
is due to ammonia. Others moisten red litmus paper with water and hang 
it in the mouth of the bottle. If much ammonia is present it will turn 
blue. (Even normal urine contains enough ammonia to turn the paper 
slightly blue in time.) 

The acidity of the urine can be increased, but not beyond a certain 
point. An increased proteid metabolism will do this or the careful adminis- 
tration of dilute mineral acids. Brown 13 has reported a series in cases 
of girls and young women of distinctly neurotic temperament with 
the urine even from 2 to 5 times the normal acidity (phenolphthalein 
as indicator) and symptoms of cystitis, i.e., pain in the trigonal region, 
but without demonstrable lesions. He suggests that it is a neurosis of 
urinary secretion. 

The urine in diabetes mellitus is very acid if it contains considerable 
oxy butyric and di acetic acids. The question of the reaction of the urine 
in the so-called " uric acid diathesis " has not yet been decided. The 
reason why it is so difficult to increase the acidity of the urine in the case 
of man is that his body protects itself against an acid intoxication by 

13 Phila. Med. Jour., March 2, 1901. 



THE URINE: GENERAL CHARACTERISTICS 103 

increasing the elimination of ammonia, thus protecting from depletion the 
native mineral alkaline store. The herbivorous animals have this ability 
to a much less degree and so they are more easily' poisoned by acids than 
is man. 

The effect of muscular work on the reaction of the urine is still doubtful. 

Some urines after standing for from 6 to 12 hours become more acid 
because of the so-called " acid fermentation." The reason for this is uncer- 
tain. It is inconstant and is always soon succeeded by an alkaline decom- 
position. Hammarsten considers it due to the reaction between the biurates 
and MH 2 P0 4 . 

Determination of the Total Acidity of the Urine. — To determine the total acidity of 
the urine Naegeli 14 added 0.1N NaOH directly to 10 c.c. of urine, using phenolphthalein 
as indicator. The error is at least from 4 to 8%. Folin 15 uses potassium oxalate in 
excess to rule out the error from ammonium salts and calcium phosphate. His method 
is as follows: 

Twenty-five cubic centimeters of urine are measured by a pipet into a 200 c.c. 
Erlenmeyer flask, I or 2 drops of 0.5% phenolphthalein solution added and 15 to 20 gms. 
of potassium oxalate. The flask is shaken well for 1 minute, then at once titrated with 
0.1N NaOH, shaking all the time. The alkali is added until a faint yet distinct colora- 
tion is produced. 

The Mineral Acidity of the Urine — Folin' s Method. — From 0.3 to 0.6 gm. of pure, dry, 
granular potassium carbonate is accurately weighed (within an accuracy of 0.2 mgm.) 
into a platinum dish and 25 c.c. of the urine to be examined measured into it. (If the 
urine contains much albumin this should be removed by heat and acetic acid. A trace 
of albumin contains too little sulphur to affect the results appreciable.) The urine is 
then evaporated on the sandbath or electric oven to dryness and when perfectly dry the 
contents of the dish are burned at just below red-heat (that is, the dish should never be 
more than faintly red-hot) over a so-called "radial burner" giving a flame wide enough 
to heat the entire bottom of the platinum dish. One must be sure the gas used does not 
contain sulphur. If there is any doubt on this point (which is tested by burning some 
of the pure potassium carbonate in the platinum dish and testing the contents for sul- 
phates) an alcohol flame may be used. If the entire bottom of the platinum dish is not 
evenly heated the cyanogen derivates of urea, which resemble mineral matter, will 
melt, flow to the cooler portions, and escape decomposition. 

The burning should continue for about an hour after all ammoniacal fumes have 
ceased to come off. Then the flame is removed. It makes little difference if the ash is 
not perfectly white. Just 10 c.c. of hydrogen peroxide water are next added, the dish 
covered with a watch glass, and gently warmed until the peroxide is decomposed. The 
watch glass is then removed and the sputterings rinsed into the dish by means of a little 
water. The contents of the dish are again evaporated to perfect dryness and are again 
heated over the radial burner as before for about an hour. The hydrogen peroxide is 
used to oxidize the thiocyanate and any small amount of sulphides which may have 
formed during the burning. Even with these precautions the complete combustion of 
the urine is very difficult. 

The residue is now dissolved in water with the help of an excess of 0.1N HC1 (75 
or 100 c.c, depending on how much carbonate was used), and is rinsed into an Erlenmeyer 
flask, boiled to drive off the carbonic acid and cooled. The excess of acid is then titrated 
with 0.1 N NaOH in the presence of a small amount of potassium oxalate (to precipitate 
the calcium) and 2 drops of a K% solution of phenolphthalein. 

14 Zeitsch. f. physiol. Chem., 1900, xxx, 313. 

15 Am. Jour. Physiol., 1903, ix, 265. 



104 CLINICAL DIAGNOSIS 

The amount of alkali and of acid added to the urine is known, one must determine: 
(i) the alkaline strength of the potassium carbonate; (2) the acidity of the hydrogen 
peroxide; (3) the SO 3 content of the hydrogen peroxide; (4) the preformed ammonia 
in the urine; (5) the inorganic SO 3 of the urine; and, finally, (6) the total S0 3 found in 
the titrated solution of the urine residue. 

The potassium carbonate and hydrogen peroxide will keep for months in well stop- 
pered glass bottles, so the first 3 determinations need be made but once (for any given 
sample of carbonate and peroxide). 

To calculate the result, one subtracts from the apparent excess of acidity found on 
titrating the burned urine residue the sum of the preformed ammonia, the acidity of the 
hydrogen peroxide and the acidity due to the organic SO3 of the urine, all in terms of 
0.1N acid. 

The acidity (in c.c. of 0.1N acid) of the organic SO 3 is obtained by subtracting the 
sum of the SO 3 of the hydrogen peroxide and the inorganic SO 3 of the urine from the 
total SO 3 of the urine residue and dividing the amount thus obtained in milligrams by 
8 (8 gms. of the organic sulphur, neutral and ethereal, are taken to represent 1 c.c. 
of 0.1N acid). 

To illustrate: 25 c.c. of urine were burned with 0.5287 gm. of potassium carbonate 
(7.76 mgms. of which contained 1 c.c. 0.1N alkali). The burned residue was boiled with 
75 c.c. of 0.1N HC1 and the titration required 1 c.c. 0.1N NaOH. An ammonia deter- 
mination gave 5.2 c.c. 0.1N NH 3 in 25 c.c. of urine. The total S0 3 = 59.9 mgms.; the 
inorganic SO 3=42.8 mgms. (10 c.c. of the hydrogen peroxide used contained 8.8 
mgms. SO 3 and 0.5 c.c. 0.1N acid). 

0.5287 gm. K2CO3 =68.1 +c.c. 0.1N NaOH 

NaOH added =i^_ c.c. 0.1N NaOH 

Total alkalinity =87.i+c.c. 0.1N NaOH 

HC1 added = 75. c.c. 0.1N NaOH 

Apparent acidity of urine = 12. 1 c.c. 0.1N HC1 

Ammonia in 25 c.c. urine = 5.2 c.c. 0.1N HC1 

Acidity of H 2 2 = 0.5 c.c. 0.1N HC1 

59.9 — (42.8+8.8) 
Acidity of organic S0 3 = « = 1. c.c. 0.1N HC1 

Mineral acidity in 25 c.c. = 12. 1 —6.7 =5.4 c.c. 0.1N HC1. 

The Organic Acidity of the Urine. — By subtracting the mineral acidity from the 
total acidity one obtains the " organic acidity," or rather the total equivalence of organic 
acid whether free or combined. In cases of acid intoxication, as in diabetes, the mineral 
acidity may turn out to be an alkalinity and all the acidity be organic. In the latter 
case the mineral alkalinity is added to the total acidity to get the organic acidity. 

THE NITROGENOUS BODIES 

The Nitrogen Output. — The total nitrogen of the urine is the best 
index of proteid metabolism. It is indeed fortunate that we have a satis- 
factory method of determining this, since it is our basal figure in all 
metabolism work. 

Folin 16 studied carefully the nitrogen distribution in the urine of 
normal men on a nitrogen-rich diet (Table I) and in 1 case on a very low 
nitrogen diet (Table II). 

16 Am. Jour, of Insanity, 1905. 



THE URINE: THE NITROGENOUS BODIES 105 

Table I Table II 

Total nitrogen 14. 8-18.2 gms. 4.8- 8.0 gms. 

Urea-nitrogen 86.3-89.4% 62.0-80.4% 

Ammonia-nitrogen 3.3- 5.1% 4.2-11.7% 

Creatinin-nitrogen 3.2- 4.5% 5.5-1 1.1 % 

Uric acid-nitrogen 0.5- 1.0% 1.2- 2.4% 

Undetermined nitrogen 2.7- 5.3% 4.8-14.6% 

Hammarsten's 17 figures are the ones quoted in most text-books. 

Normal adults On mixed diet — Infants 

Total nitrogen 10-16 gms. 

Urea 84-91 73~76 

NH 3 2-5 7.8-9.6 

Uric acid 1-3 3-8.5 

Extractives 7-12 7.3-14.7 

The sum of the nitrogens of the urea and the ammonia added together 
bears a very much more constant relationship to the total N (91 to 93%) 
than does either one alone. 

Total Nitrogen. — While the normal daily output of nitrogen is usually 
stated to be from 10 to 16 gms. since this is the average output of many 
healthy persons, yet this amount is considered by some to be evidence of 
overeating, since men can keep well and even gain weight on a diet which 
yields a daily output of but 5 or 6 gms. of nitrogen. Taylor's very careful 
work, continued over long periods of time, on the daily output of nitrogen 
in normal men, shows how wide are the variations from those limits which 
have been considered normal. Unfortunately most of the work published 
on the nitrogen of the urine is of little value since only the urea N was 
determined or due attention was not paid to the total nitrogen of the food, 
to the character of the food (its acid- or alkaline-producing qualities), 
and to the age, nutritional condition and previous diet of the patient. 
Again, the periods of observation should be at least 7 days long, during 
which time the diet and the daily amount of water consumed both should 
be constant. The patient should exercise each day a fairly constant 
amount. Granted that all these points are carefully watched, even then 
marked variations in the nitrogen output will be observed. 

By " nitrogen balance " is meant the relation of the nitrogen intake 
to the nitrogen output. The difference between these 2 figures is usu- 
ally called the " nitrogen lost " and the " nitrogen retained." When 
the output is just equal to the intake the person is said to be in 
" nitrogenous equilibrium." 

In general, the total urine nitrogen output is increased by increased 
proteid metabolism, whether by a heavy proteid diet, or by anything 
increasing the proteid catabolism of the body tissue. It is decreased by a 
diet rich in carbohydrates, in which case it reaches a lower point than dur- 
ing a fast since in the latter case the body oxidizes more of its tissue proteid. 

17 Lehrb. d. Phys. Chem., 1899, p. 421. 



106 CLINICAL DIAGNOSIS 

The total nitrogen output reaches its maximum a few hours after a heavy 
proteid meal. The evidence that exercise increases it is unsatisfactory 
since the other differences between day and night urine were not taken into 
account (see page 90) . Hot baths increase the nitrogen output. 

Any increase of water elimination will increase that of nitrogen even 
though the diet is fairly constant. One explanation given for this is that 
in the renal cells there is always a certain amount of nitrogenous waste 
which an increased water flow will wash out more thoroughly ; others say 
that the many tissue ferments follow the general law of ferments and act 
better the more dilute the solution. The output of nitrogen is diminished 
physiologically by a " poor " diet, by a reduced output of water, after 
profuse sweating, in pregnancy and after small doses of quinine. 

Pathologically the total nitrogen output is increased under the follow- 
ing conditions : in fevers, in which cases it is due not to the temperature 
per se, but, more likely, to the effect upon metabolism of the toxins causing 
the fever (the exceptions are acute nephritis causing dropsy and diseases 
with diarrhea or with the formation of large exudates) ; in diabetes, in mild 
cases if the patients are on a proteid-rich diet, but more especially in the 
severe ones who oxidize the protein of their own tissues ; after various proto- 
plasm poisons, as arsenic, antimony, phosphorus, etc.; in any condition 
which diminishes the oxygen intake, as prolonged dyspnea, hemorrhage, 
carbon monoxide poisoning, etc.; in acute lobar pneumonia during the 
resolution, that is, while there is autolysis, absorption and excretion of the 
solid exudate (in a case of Muller's the excess of nitrogen output during 
the resolution was 28 gms., which represented 800 gms. of pneumonic 
exudate) ; during the absorption of exudates or transudates ; and, finally 
by anything which increases the water output, as, for instance, diabetes 
insipidus, in which disease a daily output of 130 gms. of urea have been 
reported, and in cases of chronic nephritis with polyuria. The retention 
of nitrogen may be noted: in persons who are gaining in weight; in myx- 
edema ; during the convalescence of fevers (in a case of convalescent typhoid 
fever reported by Luthje the patient retained during 26 days 121.38 gms. 
of N, which would represent 758.6 gms. of albumin or 3568.6 gms. of 
muscle. This person gained in weight during that time 6490 gms.) ; and 
during the last stage of pregnancy. Pregnane}^ is followed by a diuresis 
and increased nitrogen output which begins about the second day of the 
puerperium. 18 The nitrogen output is diminished in all conditions which 
hinder digestion and the absorption of proteins from the intestine ; by those 
which reduce the oxidization processes in the body, as severe cachexias; 
by those conditions accompanied by large exudate and transudate forma- 
tion, as dropsy; and by renal conditions, both organic and functional, 
which hinder excretion. A marked reduction in the amount of nitrogen 
excreted is sometimes an early sign of uremia. 
18 Slemons, Johns Hopkins Hosp. Rep., 1904, vol. xii. 



THE URINE: THE NITROGENOUS BODIES 



107 



Estimation of Total Nitrogen. — Gumming s modification of the 
Kjeldahl method is quite uniformly used. From 5 to 20 c.c. of the urine, 
according to its concentration, are measured into a combustion flask of 
about 250 to 300 c.c. capacity of best quality glass and 15 c.c. of concen- 
trated sulphuric acid, 10 gms. of potassium sulphate, and about 1 gm. of 
copper sulphate are added. This flask is placed on a proper holder in a 
hood with a good draft resting on a sheet of asbestos gauze and its contents 
boiled over a free flame until clear and blue. The worker should be careful 
when washing down the carbon from the sides of the glass by shaking the 




Fig. 26. — Distilling apparatus for nitrogen determination (Kjeldahl). 

A, distillation flask; B, safety bulb; C, Liebig cooler; D, Erlenmeyer 

flask to receive distillate and containing the standard acid; E, safety 

bulb to prevent back-flow. 

fluid not to burn himself. The oxidation may be aided by adding a little 
KMn0 4 . After the fluid is perfectly blue the heat should be continued for 
a few minutes or even for half an hour, that the combustion of uric acid 
and certain other bodies may be perfect. At the end of this time practically 
all of the nitrogen will have been converted into ammonia and is therefore 
present as ammonium sulphate. The fluid is then allowed to cool, distilled 
water is next added in excess, and the fluid poured into a distilling flask 
(see Fig. 26, A) of 1 liter capacity, with long neck and round bottom. All 
of the contents of the combustion flask should be washed into this flask 
by rinsing it 3 or 4 times with distilled water. Talcum powder or zinc 
granules may now be added to prevent bumping. 



108 CLINICAL DIAGNOSIS 

That amount of strong sodium hydroxide (specific gravity 1.230) 
which has been found by previous experiments more than sufficient to 
neutralize the acid is now added and the flask at once fitted to the. Liebig 
cooler. The lower end of this cooler ends in a bent tube which descends 
vertically to the bottom of a small Erlenmeyer flask, D, of about 300 c.c. 
capacity, into which have been previously measured 50 c.c. of 0.25 N H 2 SO 4. 
In the subsequent distillation, therefore, all the ammonia, both that given 
off at once in the cold and that liberated on boiling will bubble through 
this acid and thus be caught. The distillation is continued until about 
100 c.c. of distillate have passed over. The boiling should not be too 
vigorous, and the apparatus should be protected by a safety-bulb, B, and 
watched that no acid spurt into the cooler. When the distillation is com- 
pleted the Erlenmeyer flask is lowered and the next few drops of distillate 
tested with lacmoid paper to make sure that the ammonia has entirely 
passed over. The acid clinging to the end of the tube is washed into the 
flask. This sulphuric acid is then titrated against 0.25N NaOH, using 
cochineal, methyl orange or pure litmus as indicator. Pure litmus is the 
best if the necessary precautions are used, but cochineal is sufficiently 
correct for ordinary work and is the most convenient since it can be used 
also by artificial light. (The cochineal bugs are ground fine and extracted 
with 50% alcohol. This filtered extract is used as indicator.) From 50 
c.c, the original amount 0.25N H 2 S0 4 , are subtracted the number of cubic 
centimeters of 0.25 N NaOH necessary to neutralize the acid remaining 
after the distillation. The difference is the amount of 0.2 3 N H 2 SO 4 neutral- 
ized by the ammonia. This value multiplied by 0.0035 gm. is the weight 
of nitrogen in the amount of urine used. 

(Note. — This method does not indicate nitrates or nitro compounds.) 

In many laboratories the nitrogen of the urine is determined without 
distillation. The contents of the combustion flask are poured, and the 
flask rinsed, into the apparatus for the determination of ammonia, the 
alkali added and the amount of ammonia determined (see page 123). 

The estimation of nitrogen in the urine by the Color imetric Method of 
Folin and Farmer as modified by Myers and Fine and further slightly by 
Gradwohl and Blaivas 19 requires but a few drops of urine and but from 
5 to 10 minutes of time. 

The amount of urine used should contain between 0.35 and 0.75 mgms. 
of nitrogen. One cubic centimeter of urine is measured with an Ostwald- 
Folin pipet into a 25 c.c. volumetric flask and diluted to the 25 c.c. mark 
with distilled water. (In the case of urines with a low specific gravity a 
dilution of 1 to 10 may be sufficient.) After it has been thoroughly mixed 
1 c.c. of this diluted urine is measured into a thin glass test-tube and from 
5 to 7 drops (0.1 c.c.) of concentrated sulphuric acid, 50 to 100 mgms. of 
potassium sulphate and a drop of copper sulphate (10%) added. This 

19 Jour. A. M. A., Sept. 9, 1916, vol. lxvii, p. 809. 



THE URINE: THE NITROGENOUS BODIES 109 

mixture is now boiled, while it is being shaken continuously, until it 
becomes dark brown in color and then while the tube is warm, but not 
hot, a drop of hydrogen peroxide is added and the heating continued for 
about i minute in case the fluid is not clear. The tube is now allowed 
to cool for i minute and its content then washed into a 50 c.c. volumetric 
flask (.4) with about 35 c.c. of distilled water. One now measures with 
an Ostwald-Folin pipet 5 c.c. of an ammonium sulphate solution contain- 
ing 1 mgm. of nitrogen per 5 c.c. (prepared by dissolving 0.944 gm. of 
ammonium sulphate in distilled water and making the solution up to 
1000 c.c.) into a 50 c.c. volumetric flask (B) and adds about 30 c.c. of 
distilled water. 

One now makes a fresh dilution of 10 c.c. of modified Nessler's solution. 
(This solution is made up by dissolving 100 'gins, of mercuric iodide and 
50 gms. of potassium iodide, both finely powdered, in a liter volumetric 
flask containing about 400 c.c. of distilled water. Two hundred grams of 
KOH are dissolved in 500 c.c. of distilled water, cooled thoroughly, and 
added, with constant shaking, to the mixture in the flask. This solution 
is then made up to 1 liter with water. It usually becomes perfectly clear. 
It is kept at 37 C. in an incubator over night or until the yellowish white 
precipitate which may settle out is thoroughly dissolved and only a small 
amount of dark brownish precipitate remains. The solution is now ready 
to be siphoned off for use.) Next, 10 c.c. of this solution are diluted with 
40 c.c. of distilled water, mixed thoroughly and then used to make up to 
volume the contents of the two flasks A and B. In the case of flask B, the 
Nessler's solution has neutralized the sulphuric acid. The dry, glass- 
stoppered wedge of the Hellige colorimeter is now filled with the standard 
solution and adjusted in the colorimeter. Slightly over 2 c.c. of the un- 
known solution are now poured into the empty cup, inserted in the colori- 
meter, and the colors matched, preferably by a North light. The amount 
of nitrogen in %5 c.c. of urine, the amounts actually used, may be ascer- 
tained from Table I (page no). 

Since the figures in the table are given for dilution of 100 c.c. and the 
dilution here employed is 50 c.c, the result obtained should be divided 
by 2. 

Urea is the nitrogenous body of the urine present in largest amount 
and the one which until recently has attracted most attention. Since none 
of the methods formerly employed to determine it 20 gave reliable results, 
the great mass of research work done using these methods must be con- 
sidered inaccurate. 

The output of urea has been used as a test of the digestion. A meal 
containing an excess of nitrogen is ingested; for illustration, 500 gms. of 
meat, 8 eggs and 200 gms. of bread. During this and the following day 
at least 50 gms. of urea should be excreted. 

20 Liebig's, Hiifner's, Moerner-Sjoquist, Schoendorf, Folin's, et al. 



110 



CLINICAL DIAGNOSIS 

TABLE I * 



Estimation of Nitrogen with the Hellige Colorimeter 



Calorimetric 
reading 


Nitrogen mgms. 

per dilution of 

100 c.c. 


Colorimetric 
reading 


Nitrogen mgms. 

per dilution of 

100 c.c. 


Colorimetric 
reading 


Nitrogen mgms. 

per dilution of 

100 c.c. 


20 
21 

22 

23 
24 


i-73 
1.71 
1.69 
1.67 
1.65 


40 

41 

42 

43 

44 


1 .31 

I.29 
1.27 

1-25 

1.23 


60 
61 
62 

63 
64 


O.89 

O.87 
O.85 
O.83 
O.81 


^5 
26 
27 
28 
29 


1.62 
1.60 
1.58 
1.56 
1-54 


45 
46 

47 
48 
49 


1.20 
1. 18 
1. 16 

1. 14 
1. 12 


65 
66 

67 
68 
69 


O.78 
O.76 
O.74 
O.72 
O.70 


30 
31 
32 

33 
34 


1.52 
1.50 
1.48 
1.46 
1.44 


50 
5i 
52 
53 
54 


1. 10 
I.08 
1.06 
I.O4 
I.02 


70 

71 

72 

73 
74 


O.67 
O.65 
O.63 
0.61 
0.59 


35 
36 

37 
38 
39 


1.41 
i-39 
1-37 
i-35 
i-33 


55 
56 

57 
58 
59 


O.99 
O.97 
0.95 

0-93 
0.91 


75 
76 
77 
78 
79 


O.56 

0-54 
O.52 
O.50 
O.48 



* Myers and Fine's table copied from Gradwohl and Blaivas. 

The normal person on an average diet is said to excrete from 20 to 40 
gms. of urea each 24 hours; on a poor diet, only 15 or 20 gms. ; if on a very 
rich diet, even 100 gms. in 24 hours. Men are said to eliminate more than 
women. In general it may be said that a vigorous person will eliminate 
on an average diet about 30 gms. and an invalid on a liquid diet about 
20 gms. per day. 

The amount of urea eliminated may be diminished because the output 
of total nitrogen is diminished or because the nitrogen is excreted in some 
other form than urea, e.g., as ammonia. One of the most important func- 
tions of the liver is to change the ammonia bodies to urea ; hence in certain 
diseases which decrease the liver function the output of urea will diminish 
and that of ammonia increase until even from 50 to 60% of the total 
nitrogen is eliminated as ammonia (see page 122). It is also true, however, 
that some cases with marked gross lesion of the liver will eliminate urea 
and ammonia in normal percentages. Again, an unusually large per cent, 
of the nitrogen may be eliminated as ammonia because of acids which are 
ingested or formed within the body. These must be neutralized and to 
protect the mineral alkali of the blood and lymph, nitrogen "which other- 
wise would, appear as urea will be eliminated as ammonia and so be with- 
drawn from urea formation. This occurs in diabetes and in cachexias 
which disturb the absorption or use of carbohydrates. 



THE URINE: THE NITROGENOUS BODIES 111 

The interesting question has been raised, Why is there any great excess 
of urea at all in the urine? There are various answers to this question. 
One is that urea is the chief nitrogenous ash of nitrogenous food and that 
a normal American on an " average " American diet should excrete from 
20 to 30 gms. of it each day. Another opinion is that the urea represents 
that part of our nitrogenous intake which is over and above that which 
we really needed and that man " living rationally " would have very little 
urea in his urine. Another view, not so radical, is that not all of the split 
products of protein digestion are resynthesized to protein ; that the cleavage 
liberates the carbonaceous portion of the protein molecule and the various 
amido acids, some of which are needed then, others not, and that to get a 
sufficient amount of a great enough variety of these amidobodies a great 
deal of proteid must be torn down, the most of which must be rejected ; that 
it is this nitrogen which is finally excreted as urea. It is, however, quite 
definitely established that urea is the on]y nitrogenous ash which is dimin- 
ished both absolutely and relatively when the total nitrogen output is 
diminished and that a man can keep in nitrogenous equilibrium and in 
good ( ?) health for a limited time on an astonishingly limited diet (see the 
table on page 105). 

Urea, when pure, crystallizes out in needles or prisms belonging to the 
tetragonal system, which are colorless, striated, pale, four-sided columns 
with ends in 1 or 2 oblique planes and which sometimes are hollow. They 
contain no water of crystallization, are not hydroscopic, and do not 
change in the air. They are decomposed by heat, liberating ammonia, 
the decomposition beginning at ioo° C. and becoming most active at 
130° C. 

The furfurol test, the most important for urea, is made, according to 
Schiff, by bringing 1 crystal of urea the size of the head of a pin in contact 
on a porcelain dish with 1 drop of concentrated aqueous solution of fur- 
furol and adding at once 1 drop of hydrochloric acid (specific gravity, 
1.1). A rapid change of colors takes place; first yellow, then green, blue, 
violet, and in a few minutes a fine purple- violet. Alantoin gives the same 
test but less intensely and more slowly. An old furfurol solution should 
not be used since it may give this change of colors even in the absence of 
urea. Huppert mixes 2 c.c. of concentrated furfurol solution with 4 to 6 
drops of concentrated hydrochloric acid. If no red color is produced, 1 
crystal of urea is added. In a few minutes the fluid becomes a deep violet 
color, which gradually turns to black and a black precipitate forms. 

The biuret test is 1 of the best-known tests for urea. Urea if fused at a 
temperature of ioo° C. gives off biuret and cyanuric acid. To make this 
test a few crystals of urea in a dry test-tube are heated gently until fluid, 
then cooled, dissolved in water, made strongly alkaline with NaOH, and 
then a 2% solution of Q1SO4 added drop by drop. A beautiful violet 
color will be the result. 



112 CLINICAL DIAGNOSIS 

If one has but a minute quantity of material at his disposal, e.g., a 
grain of skin frost, the best tests for urea are the nitric acid or oxalic acid 
tests. One crystal or i drop of the concentrated solution (at least 10%) 
is allowed to come in contact under the cover-glass with pure nitric acid. 
At the line of contact the characteristic crystals of urea nitrate CO(NH 2 )2 
HN0 3 form rapidly as colorless rhombs or hexagonal plates with acute 
angles which often overlap like shingles. If they crystallize out slowly 
it is in the form of large, thick, rhombic prisms. These crystals, when 
heated, volatilize without leaving any residue, an essential point to exclude 
similarly shaped crystals of the heavy metals. The nitric acid used should 
be free from nitrous acid which would decompose the urea forming carbon 
dioxide, nitrogen, and water. 

In a similar test oxalic acid is used instead of nitric acid. The urea 
oxalate, 2CO(NH 2 ) 2H2C2O4 formed is less soluble in water than the nitrate, 
which is an advantage. These crystals are rhombs, hexagons, or plates. 

A still better method is to dissolve the urea in the least possible amount 
of absolute alcohol and to bring this in contact with a concentrated 
ether solution of oxalic acid or, better still, to use an amyl alcohol 
solution of both. 

Quantitative Determinations of Urea — Urease Method of Marshall. 
Into each of two 200 c.c. Erlenmeyer flasks are measured 1 or 2 c.c. of 
toluol. Into 1 is measured exactly 5 c.c. of the urine, the urea of which 
is to be determined, and 100 c.c. of distilled water. One urease tablet is 
crushed in a glass mortar, dissolved in about 5 c.c. of water and washed 
into the second flask using for this purpose in all about 90 c.c. of water. 
Five cubic centimeters of the urine are then carefully measured into this 
flask. Both flasks are now tightly closed with corks and their contents 
agitated. They are then allowed to stand at room temperature for at 
least 8 hours. If haste is desired 2 tablets may be used instead of 1 
and the flasks incubated in the thermostat at 40 C. for 1 hour; or, but 
1 c.c. of urine may be used, 2 tablets of urease, 100 c.c. of distilled water 
and the flasks digested at from 40 to 50 C. for 15 minutes only. 

After the proper time of incubation has elapsed the contents of both 
are titrated with 0.1N HC1, methyl orange used as indicator, until they 
assume a distinct pink color. The amount of 0.1N HC1 required to neu- 
tralize the specimen containing the urease less the amount required, to 
neutralize the control (the preformed ammonia) will give the urea content 
of 5 c.c. of urine estimated as ammonium carbonate. One cubic centimeter 
of 0.1N HC1 will indicate 0.001401 gm. of N and this value multiplied by 
2.143 the amount of urea. 

Quantitative Determination of Urea by the Urease and Colorimetric 
Method of Folin. — By this method the urea is converted by the urease into 
ammonium carbonate, the ammonia then liberated by sodium carbonate 
in excess and drawn over by aeration into hydrochloric acid. The ammon- 



THE URINE: THE NITROGENOUS BODIES 



113 



ium chloride formed can be determined colorimetrically by the use of 
Nessler's reagent. 

The urine is diluted i to 10 with distilled water, 2 c.c. of this measured 
into a test-tube of such dimensions that it will easily slip into a 100 c.c. 
narrow cylinder (Fig. 27, B) without lip, about 0.1 gm. of urease added 
and the contents then incubated for l / 2 an hour in a beaker of water at 50 C. 
Two drops of caprylic alcohol or 1 c.c. of amy lie alcohol are next added to 
prevent foaming during aeration. 

The apparatus for aeration consists of two 100 c.c. cylinders for each 
sample of urine. If more than 1 specimen is to be examined, and control 
determinations always should be made, the 4 cylinders may be run in series. 
Of the 2 cylinders for each test the 1 is graduated, the other not graduated 
and both are provided with 2 -hole rubber stoppers. Cylinder A is graduated 
and is connected by tube a with the suction apparatus. Cylinder B 

a' 



^w^m 



w 












e 



Fig. 27. — Showing the urea apparatus set up and connected to suction. 



is not graduated and is connected with the acid wash bottle (C) . If more 
than 1 urine is under examination, cylinder B is connected with the short 
connection tube of the other graduated cylinder A. The wash-bottle C 
contains sulphuric acid (10%) and is placed at the end of the outfit to 
prevent any ammonia in the air from gaining entrance into the system. 
Tube a is bent at right-angles and extends only to a point just within 
cylinder A . The tube b which extends almost to the bottom of cylinder A 
ends in a bulb pierced by a number of small holes (made with a platinum 
wire at white heat while the glass is only moderately hot). Cylinder B 
is connected with A by a right-angle tube extending to a point just below 
the stopper. Its other tube c has a straight open end long enough to dip 
into the test-tube (E) while its other end is bent at right angles and serves 
as connection either with the acid wash bottle or with the other series of 
cylinders in case more than 1 urine is to be examined. Into cylinder^, are 
measured 20 c.c. of distilled water and 2 to 3 drops of 10% hydrochloric 
acid. This is now closed and cylinder B opened. To the digested urine in 
the test-tube an equal volume of saturated sodium carbonate is added, 
8 



114 CLINICAL DIAGNOSIS 

this being allowed to run slowly down the side of the tube under the urine, 
the tube is now quickly and carefully placed in cylinder B which is then 
at once closed, care being taken that tube c reach almost to the bottom 
of the test-tube, and all the connections carefully, sealed. The suction, by 
means of the Chapman pump, is started slowly for about 5 minutes, then 
increased to the limit and continued for from 30 to 45 minutes. The 
stopper of c}dinder A is now removed, care being taken to wash back all 
fluid on tube b with 2 to 3 c.c. of distilled water. 

Into a 50 c.c. volumetric flask is pipeted 5 c.c. of an ammonium sul- 
phate solution 5 c.c. of which contain 1 mgm. of nitrogen (see page 109), 
25 c.c. of distilled water and 20 c.c. of Nessler's solution (see page 109) 
diluted 1 to 5 . To cylinder A , which contains the nitrogen of the urea in 
the form of ammonium chloride, is added from 10 to 2 5 c.c. of diluted (1 to 5) 
Nessler's solution, the amount depending upon the depth of color and 
this then diluted to 50 c.c, 100 c.c, etc (depending upon the color). The 
colorimetric reading should be made at once with the Hellige colori- 
meter (see page 109) and the result calculated with the aid of Table I. 

The result will be the amount of nitrogen in 0.2 c.c. of urine (the urine 
was diluted 1 to 10 for this test and 2 c.c of diluted urine taken for 
the determination). 

Suppose the reading was 58. According to Table I, this would indicate 
0.93 mgm. per 100 c.c. of a dilution which contained 0.2 c.c. of urine. This 
multiplied by 5 gives the amount of nitrogen in t c.c. of urine; from this 
should be subtracted the amount of ammonia nitrogen originally present 
and determined separately. The difference multiplied by the factor 2.14 
gives the amount of urea which would contain that amount of nitrogen. 

To isolate urea from any solution, all albumin should first be removed. 
Then the solution, faintly acidified if necessary, is concentrated at a low 
temperature to a very small volume. Nitric acid is then added in excess, 
the mixture meanwhile being kept cool. The precipitate is filtered and 
pressed between filter paper. It is then dissolved in water, decomposed 
with barium carbonate, dried upon a water-bath and the residue extracted 
with strong alcohol. The extract is decolorized if necessary with animal 
charcoal. When this is cooled urea will crystallize out from the warm alco- 
holic solution. 

Uric acid is a substance which has attracted an absurd amount of atten- 
tion and been the object of a vast amount of time-consuming work. The 
present consensus of opinion is that it is a specific oxidization product of 
the nuclein basis and is increased in the urine only as a result of an increase 
of these bodies in the food, or of an increased metabolism of the tissue 
nuclei. Horbaczewski believed that it is derived especially from the nuclei 
of leucocytes. This probably explains but a small fraction of it. It is 
interesting that in the excrement of birds and certain reptiles uric acid 
is the most important nitrogenous substance while in that of some carniv- 



THE URINE: THE NITROGENOUS BODIES 115 

ora (dogs and cats) little or none can be demonstrated. In the urine of 
herbivora traces of it are constantly present, while in man it is excreted 
in a fairly large but still very varying amount. It has been shown that 
our body has the ability on the one hand to oxidize uric acid and on the other 
to synthesize it. If hypoxanthin, e.g., be fed a patient, 50% will appear 
as uric acid. It is probable that in birds, as in mammals, urea is the chief 
end product of nitrogen metabolism but that their tissues synthesize it 
further to uric acid, while in mammals it is excreted unchanged. Not- 
withstanding these opportunities for fluctuation in the amount excreted 
the recent work tends to prove that uric acid is a very specific product of 
the oxidization of nuclein bases and that variations in its output would 
seem to be due in large measure to delays in its elimination. 

Uric acid, when pure, is a white crystalline powder consisting of very 
small prisms or plates. It is with difficulty soluble in boiling water and 
very little in cold. It is more soluble if not pure. Urea is its best solvent 
and explains its presence in solution in the urine. It is insoluble in alcohol 
and ether; is somewhat soluble in hydrochloric acid and in solutions of 
the alkaline carbonates. The cold solution of uric acid does not redden 
litmus. It reduces Fehling's solutions when heated, but not Nylander's 
solution (see page 170). It is decomposed by NaOBr which will liberate 
about 47.8% of its nitrogen. 

The normal output of uric acid in the urine of man varies from 0.2 to 
1.25 gms., an average of 0.7 gm. in 24 hours. This represents from 0.5 to 
2% of the total nitrogen of the urine. It is increased physiologically by 
increasing the nucleins of the food. A meal of sweetbreads, e.g., will cause 
an increase in its output of from 0.5 to 2 gms. in 24 hours. The maximum 
output occurs from 3 to 5 hours after a meal (that of the nitrogen in 9 
hours). There is a relatively large output in the urine of the new-born. 
In the adult the nitrogen of the uric acid is to the nitrogen of the urea as 
1 : 50 to 70, but in the case of the new-born as 1 : 13 to 14. 

The amount excreted by a normal person on a mixed diet varies con- 
siderably from day to day and the differences between different persons are 
considerable. Burian and Schur have simplified this problem greatly by 
showing that the uric acid output may be divided into 2 fractions — the 
exogenous and the endogenous. By exogenous is meant the uric acid which 
is formed from the food directly ; the endogenous, that part arising from the 
tissue proteid. The endogenous fraction is therefore the more interesting 
fraction to consider and in metabolism work involving uric acid the patient 
should be on a diet, e.g., of eggs and milk, which furnishes sufficient nitrogen 
and heat but contains no nucleins. Although for this reason most of the 
quantitative work on uric acid must be discarded }^et it is agreed that uric 
acid is pathologically increased when there is an increased proteid catabol- 
ism, as in fevers in which cases the increase of uric acid runs parallel to 
that of the urea. There is an absolute increase in leukemia, the record 



116 CLINICAL DIAGNOSIS 

output being 8 gms. in 24 hours (reported by Magnus-Leyy ) , although in 
most cases of this disease it is about 2 gms. per day and its nitrogen is to 
that of the urea as 1 : 9 (normal 1 : 50-70). While the importance of uric 
acid in gout is still uncertain, it would seem to be true that between the 
attacks its output is below normal and that it rises to normal with the 
acute symptoms. This may help in the diagnosis of a doubtful case of 
arthritis. The explanation of this would seem to be a retardation in the 
formation and excretion of uric acid although the large accumulations of 
biurates in the tophi and around the joints are usually cited as evidence of 
an increased uric acid production. In other forms of arthritis the question 
is still unsettled. In diabetes mellitus the increase in uric urine is not 
marked, only to 2 to 3 gms. per day, and is due to the diet; in pernicious 
anemia an increase is claimed. In pneumonia during resolution the output 
is increased, probably because of the breaking down of the nuclei of the 
cells of the exudate. In cirrhosis of the liver its output is said to be very 
much increased, Chabria claiming in certain cases even 8 gms. in 24 hours. 
This is interesting since the liver certainly can synthesize uric acid. The 
much discussed uric acid diathesis theory so emphasized by Haig and others 
is still in doubt and is losing ground. 

The output of uric acid has been found diminished by a poor diet, in 
nephritis, during the acute attack of gout, in certain chronic diseases and 
after large doses of quinine. 

It is of interest that when the alloxuric bases are increased in the urine 
the uric acid decreases in the same proportion. 

The urates described are: 

(1) Neutral, MU, which are not found in nature. 

(2) The monoacid- or biurates, MHU, which are gelatinous or crystal- 
line bodies, the best illustration of which are the needles found in 
gouty tophi. 

(3) Quadriurates, MHUU (Roberts), which are easily split to MHU 
and U by water, heat or acid. They are less soluble than the biurates. 
They are supposed to make up the common urate sediment. Many 
observers think the so-called quadriurates are merely mixtures of sodium 
biurate and uric acid. 

The murexid test for uric acid is the one in common use. The crystal to 
be tested is dissolved in 2 drops of nitric acid and evaporated carefully 
to dryness. The residue will have a beautiful red color. Ammonia is then 
added, whereupon the color changes to a purple red. Had NaOH or KOH 
been used in place of the NH 4 OH, the color would be more blue and this 
is an important point in excluding certain other bodies. This test is more 
brilliant if the nitric acid is evaporated over a water-bath and if the am- 
monia, not added directly, is placed in a small glass under a bell- jar near 
that containing the dried residue ; also if but little uric acid is used. If the 
residue is not red but yellow too little nitric acid was used and more should 



THE URINE: THE NITROGENOUS BODIES 117 

be added and the evaporation repeated. It is an essential part of this 
test to bleach this color by heat. 

Guanin, xanthin, epiguanin, also will give this test, but these are ex- 
cluded if the substance used was insoluble in an excess of HC1. 

In addition to a positive murexid test the ability of the substance in 
question to reduce Fehling's solution should be tested. 

Quantitative Determination of Uric Acid. — Folin's Method. — To 
300 c.c. of urine are added 75 c.c. of an uranium acetate reagent (consisting 
of 500 gms. of ammonium sulphate and 5 gms. of uranium acetate dissolved 
in 650 c.c. of water; 60 c.c. of 10% acetic acid are then added and the 
volume made up to 1 liter. This solution is to remove the phosphates and 
certain bodies not well understood whose presence in certain pathological 
cases disturbs the accuracy of the method) . The urine thus treated is well 
stirred, allowed to stand for 5 minutes and filtered through a double folded 
filter. Of this filtrate 125 c.c. (representing therefore 100 c.c. of urine) are 
measured into each of 2 beakers, 5 c.c. of concentrated ammonia are added 
to each and the beakers set aside until the next day to allow the precipitate 
of ammonium urate to settle. The .clear fluid is then decanted through a 
filter paper and the precipitate finally collected on this paper and washed 
with a 10% solution of ammonium sulphate until the filtrate is almost 
chlorine-free. (This is tested by adding to a little of the filtrate HNO3 
till it is strongly acid and then a drop of AgN0 3 .) The filter paper is then 
pierced and the ammonium urate washed into a beaker, using for this 
purpose about 100 c.c. of water. Fifteen cubic centimeters of concentrated 
sulphuric acid are then added and the solution titrated while still hot 
with 0.0 5 N KMn0 4 solution until the first blush of red persists for a few 
seconds throughout the whole volume of fluid. Each cubic centimeter of 
the reagent used indicates 3.75 mg. of uric acid. It is necessary to add as a 
correction 3 mg. of uric acid per 100 c.c. of urine. 

A 0.05N KMn0 4 solution is one of such concentration that 1 liter would 
contain 0.05 gm. of available oxygen with which to oxidize the uric acid. 
vSuch a solution would contain therefore 1 .581 gms. of recrystallized KMn0 4 
in 1 liter of water. Since KMn0 4 cannot be weighed with sufficient 
accuracy it is best to make a slightly more concentrated solution, to boil 
this, which renders it more permanent and then titrate it against a 0.1N 
solution of oxalic acid (6.3 gms. per liter) or 1 of potassium tetraoxalate 
(8.41 gms. per liter). Ten cubic centimeters of this oxalic acid solution are 
diluted to 100 c.c. with distilled water, 15 c.c. of concentrated sulphuric 
acid are then added, which will produce a temperature of about 6o° C. 
and the potassium permanganate solution added drop by drop from a 
buret until a uniform red color remains about 30 seconds throughout the 
entire volume of fluid. The permanganate solution is then so diluted that 
10 c.c. of the oxalic acid will require 20 c.c. of the KMn0 4 solution to 
produce this end reaction. 



118 CLINICAL DIAGNOSIS 

It is interesting that at the beginning of the titration of uric acid the 
red color remains longer than later. This is due to the fact that the com- 
bustion of the uric acid is much promoted by the increasing percentage of 
the sulphate of manganese. The color is not permanent owing to the 
presence of other reducing bodies in the urine and the student, to use the 
test satisfactorily, should standardize his own solutions, that he may know 
what to consider the end reaction. 

To obtain oxalic acid sufficiently pure it is necessary to recrystallize 
it 2 or 3 times from a cold saturated solution; or, better, to recrystallize 
it first from hot dilute HO (io to 15%), then from hot alcohol and then 
from water. The aqueous solution must be heated till the odor of ethyl 
oxalate has passed off. Oxalic acid cannot be dried in a desiccator or on a 
hot-air bath. 

If from the urine to be examined some uric acid or urates have already 
precipitated, this sediment should be redissolved by warming the urine, 
or by the addition of a little saturated lithium carbonate solution and the 
urine well shaken before it is used. 

Colorimetrical Method of Determination. — Into a 15 c.c. conical, cen- 
trifuge tube one pipets 2 c.c. of urine, adds 15 drops of ammoniacal- 
silver magnesium mixture, inverts the centrifuge tube in order to mix 
its contents and then allows it to stand for about 10 minutes in a refrig- 
erator. At the end of this time the tube is centrifugalized for from 3 to 5 
minutes and the supernatant fluid poured off by inverting the tube and 
wiping its lip with filter paper. The ammonia is now removed from the 
precipitate by volatilization by attaching the mouth of the tube to the 
suction apparatus. 

From this point on the student must work as fast as possible as the 
colors may fade or the solution become turbid. 

One now makes up a standard solution in a 50 c.c. volumetric flask by 
pipeting into it 5 c.c. of standard uric acid solution (5 c.c. of which equals 

1 mgm. of uric acid) adds 2 drops of a 5% solution of potassium cyanide, 

2 c.c. of Folin-Macallum reagent, 20 c.c. of saturated sodium carbonate 
and in 1 minute fills up with water to the 50 c.c. mark. 

To the precipitate in the centrifuge tube (which is now free from 
ammonia) are added 2 drops of a 5% solution of potassium cyanide, the 
tube shaken so as to dissolve the precipitate, then 2 c.c. of Folin-Macallum 
reagent and the contents of the tube washed into the 100 c.c. graduate 
with from 15 to 20 c.c. of saturated sodium carbonate solution (with 20 
c.c. if the color is well developed, 15 c.c, if fainter). Since it is quite 
important that this, the unknown solution, be weaker in color than the 
standard, one now waits for from 40 to 60 seconds before determining from 
the depth of color whether to dilute it to 50 c.c. or 100 c.c. 

The readings are then made with the Hellige colorimeter using Table II 
to estimate the amount of uric acid present. 



THE URINE: THE NITROGENOUS BODIES 119 

TABLE II * 
Estimation of Uric Acid with Hellige Colorimeter 



Calorimetric 
reading 


Uric acid mgms. 

per dilution of 

100 c.c. 


Calorimetric 
reading 


Uric acid mgms. 

per dilution of 

100 c.c. 


Calorimetric 
reading 


Uric acid mgms. 

per dilution of 

100 c.c. 


20 
21 

22 
23 

24 


I.67 
1.65 
I.63 
I.6l 
1-59 


40 
41 

42 

43 
44 


1.28 
1.26 
I.24 
1.22 
I.20 


60 
61 
62 

63 
64 


O.88 
O.86 
O.84 
O.82 
0.8l 


25 
26 

27 

28 

29 


i-57 
i-55 
1-53 
1.51 
1.49 


45 
46 

47 
48 
49 


1. 18 
1. 16 
1. 14 
1. 12 
I. IO 


65 
66 

67 
68 
69 


O.79 
O.77 
0-75 
0-73 
O.71 


30 
3i 

32 

33 
34 


1.48 
1.46 
1.44 
1.42 
1.40 


50 
5i 
52 
53 
54 


1.08 
1.06 

I.04 
1.02 
1. 00 


70 
71 

72 

73 
74 


O.69 
O.67 
O.65 
O.63 
0.6l 


35 
36 
37 
38 
39 


1.38 
1.36 
1-34 
1.32 
1.30 


55 
56 
57 
58 
59 


O.98 
O.96 
O.94 
O.92 
O.9O 


75 
76 
77 
78 
79 


0-59 
0.57 
0.55 
0.53 
O.51 



* Myers and Fine's table copied from Gradwohl and Blaivas. 

Example 1. — Suppose the dilution is to 100 c.c. and the reading 60. 
The equivalent of 60 as given in the table is 0.88 mgm. in the amount of 
urine which was diluted to 100 c.c, i.e., in 2 c.c. of urine. One cubic centi- 
meter of the urine contained therefore 0.44 mgms. of uric acid, and since 
uric acid contains 33% nitrogen, 0.1452 mgms. of N. 

The ammoniacal-silver magnesium mixture is made up by mixing 70 c.c. 
of 3% silver nitrate solution, 30 c.c. of magnesium mixture and 100 c.c. 
of concentrated ammonia. Any turbidity which may develop is removed 
by nitration. 

This magnesia mixture is made by dissolving 35 gms. of magnesium 
sulphate and 70 gms. of ammonium chloride in 280 c.c. of distilled water 
and then adding 140 c.c. of concentrated ammonia. 

For the preparation of uric acid standard solution, one dissolves 9 gms. 
of pure crystalline hydrogen disodium phosphate and 1 gm. of dihydrogen 
sodium phosphate in from 200 c.c. to 300 c.c. of distilled water, filters and 
makes up to about 500 c.c. with hot distilled water. This warm, clear 
solution is now poured on 200 mgms. of pure uric acid (Kahlbaum) sus- 
pended in a few cubic centimeters of water in a liter flask. The flask is 
agitated until the uric acid is completely dissolved and exactly 1.4 c.c. 
glacial acetic acid at once added, the solution made up to 1 liter, mixed, 
and 5 c.c. of chloroform added. Five cubic centimeters of this solution 



120 CLINICAL DIAGNOSIS . 

are equivalent to i mgm. of uric acid. This solution should be freshly 
prepared every 2 months. Before weighing out the 200 mgms. of uric 
acid, it is well to dry the bulk of the uric acid from which the above amount 
is to be weighed in a drying oven overnight at ioo°. 

For the preparation of the Folin-Macallum reagent, one boils 100 gms. 
of sodium tungstate, 20 c.c. of concentrated hydrochloric acid and 30 c.c. 
of 85% phosphoric acid in 750 c.c. of distilled water for 2 hours and then 
makes it up to 1000 c.c. During the boiling it is well to have a funnel over 
the flask so as to prevent undue evaporation. 

Rudisch and Kleeberg 21 have reported a method for determining uric acid and the 
purin bases which they think superior in accuracy even to the Ludwig-Salkowski method 
and so simple that it can be used clinically. They precipitate all these by an excess of 
0.02N AgNCh, and then determine the excess of silver volumetrically by titration with 
0.02N KI. The end reaction is recognized by testing the mixture in test-tubes after 
the addition of each successive portion of KI with nitrous-sulphuric acid (25 c.c. H2SO4 
to 75 c.c. H 2 0, then 1 c.c. of fuming HN0 3 ) and starch solution, until the blue of starch- 
iodine compound appears. The separation of uric acid from the other purin bodies 
depends on the solubility of the silver compounds of the latter in strong ammonia 
solutions. 

The Purin Bases. — The purin, alloxuric, xanthin, or nuclein bodies, as 
they have been called, are found in the urine in very small amounts. These 
bodies all are compounds of the purin nucleus combined with the amido, 
oxy, and methyl groups. Of those present in the urine some are exogenous 
that is, are derived wholly from food, but others are of endogenous origin, 
i.e., are end products of the clearage of the tissue nucleins. Their formation 
is represented by the following diagram : 

Nulcein 

Simple proteid Nucleic acid 

Thymic acid Purin bodies 

Thymin Metaphosphoric acid 

Uracil 

Cytosin 

'ilie most important of the purin bodies are xanthin and hypoxanthin. 
Among the others are adenin, episarkin, and epiguanin. Some have claimed 
to have found guanin and carnin in the normal urine but that is as yet 
unconfirmed. That amount of each which appears in the urine is the residue 
which has escaped transformation to uric acid. The 3 which make up the 
bulk of the purin bodies found in the urine are heteroxanthin, paraxanthin 
and methylxanthin, and these are exogenous in origin, that is, are derived 
directly and wholly from the caffeine, theobromine, and theophylin of the 
food. The total amount of purin bodies found in the urine varies from 
15.6 to 45.7 mgms. in 24 hours. Others consider that, for a mixed diet, 87 



21 Am. Jour, of Med. Sci., 1904, vol. cxxviii, p. 899. 



THE URINE: THE NITROGENOUS BODIES 121 

mgms. is an average output (Camerer), 44 mgms. that for a meat and in 
mgms. for a vegetable diet. 

Xanthin occurs normally in the urine in minute traces. From 10,000 
liters of normal urine 16 gms. of xanthin have been isolated. Very rarely 
it is the chief constituent of a urinary sediment or even of a calculus, 
several of which have been described. It is increased in leukemia, in the 
nephritis of children (in which cases even 28.5 mgms. per 100 c.c. instead 
of, as normally, 3.8 mgms. have been reported). 

The principal test of xanthin is WeideVs Test. The substance in ques- 
tion is boiled in a test-tube with hydrochloric acid and a little KC10 3 . 
It is then carefully evaporated to dryness, and the residue moistened with 
ammonia. A red or a purple-violet color results. Another test is to add 
HNO3 and evaporate to dryness in a porcelain dish, which will produce a 
yellow residue. On the addition of NaOH and warming, this becomes a 
purple-red color. 

Guanin is said to have been found in the urine, especially in leukemia. 
It gives the same test with nitric acid as xanthin, excepting that the addi- 
tion of the alkali produces a more blue-violet color. It does not give 
the Weidel reaction. 

Hypoxanthin is present in the normal urine and in considerable 
amounts in leukemia. It gives neither the nitric acid nor the Weidel tests. 

Adenin occurs in urine, especially in leukemia. The characteristic 
reaction of this substance is that if its crystals be warmed slowly in an 
amount of water insufficient to dissolve them, when the temperature reaches 
50 C. there will appear a sudden cloud. It does not give the nitric acid 
nor the Weidel test. Its other reactions are the same as hypoxanthin. 

The best method of the quantitative determination of these bodies is that of 
Salowski. From 400 to 600 c.c. of urine (albumin removed) are precipitated by a mag- 
nesium mixture and filtered. The filtrate is then precipitated with a 3% ammoniacal 
silver solution (6 c.c. per ioo c.c. of urine) and filtered. This silver precipitate is washed 
thoroughly and then suspended in about 600 to 800 c.c. of water, slightly acidified with 
hydrochloric acid and decomposed with H 2 S. The fluid is then heated to boiling and 
filtered hot. The filtrate is evaporated on a bath to dryness and the residue extracted 
with 3% hot sulphuric acid, from 25 to 30 c.c. being used. The extract is allowed to 
stand for 24 hours. 

The uric acid is then filtered out and washed, the filtrate made alkaline and again 
precipitated with AgN0 3 . This precipitate is then collected on a small chlorine-free 
filter, washed, dried and carefully ashed, the ash dissolved in nitric acid and titrated 
for chlorides by the ordinary Volhardt method. One part of silver equals 0.277 parts 
of the xanthin base nitrogen, or 0.7381 parts of the xanthin bases. The uric acid can 
be determined in the same portion. 

The enormous literature on the xanthin bases has lost its value since 
some of the methods used have been found incorrect. It is quite certain, 
however, that in leukemia these bodies are increased, also in tuberculosis ; 
and that their output bears a reciprocal relation to that of uric acid . 



122 CLINICAL DIAGNOSIS 

Ammonia. — The figures usually given for the 24-hour output of ammonia 
in normal urine (from 0.3 to 1.2 gms., average 0.7 gm.) are considered by 
Taylor to be much too high. He found that if the urine be carefully pro- 
tected from all decomposition only about Xo of the above amount will be 
present. Under normal conditions the elimination of ammonia reaches its 
maximum during sleep — that is, when digestion is at rest. Many believe 
that the ammonia in norma] urine is that which has been withheld 
from urea formation in order to balance acid ions. But this theory 
cannot explain all, for some will be present even after long continued 
alkaline medication. 

Ammonia is one of the most important end-products of proteid metab- 
olism. In the arterial blood there is 0.4 mgm., and in the portal blood 1.85 
mgm. in 100 c.c. (Hordynski). It is found in all the tissues, especially in 
the stomach wall which contains 36.4 mgms., and the intestinal wall which 
contains 32.4 mgms. per 100 gms. of these tissues. It is especially abundant 
in these organs at the height of digestion. In the other organs the amount 
is more constant. Under normal conditions the ammonia bodies, all of 
which are rather toxic, are rapidly synthesized to urea, by the liver especi- 
ally, therefore in certain hepatic diseases — e.g., in far-advanced cirrhosis 
and cancer — while the total nitrogen output remains unchanged the per- 
centage of urea tends to fall and that of ammonia to rise. 

The relation of N : NH 3 is quite constant if the diet is constant and is 
not affected by the amount of proteid consumed. If much fat be added, 
however, the percentage of NH 3 is increased. During the secretion of the 
HC1 of the gastric juice the percentage of nitrogen in the urine rises. 

The ammonia of the urine is increased: by the ingestion of inorganic 
acids and of organic acids which cannot be further oxidized, as well as by 
any increase in the amount of acids which arise in the body and they do 
this by using ammonia as a base before it can be changed to urea. Man and 
the carnivora, it may be said with truth, are constantly defending them- 
selves against an excess of acid radicals produced by the catabolism of 
animal proteids and they do this by using ammonia as a base, thus protecting 
their native alkalinity from depletion. The herbivora protect themselves 
less well than man and so suffer more quickly. It is increased by a diet 
rich in protein, or fat; in conditions producing oxygen starvation; in fever, 
during the febrile stage and continuing into the convalescence (Rumpf) ; 
in diabetes mellitus, in which cases oxy butyric and diacetic may be demon- 
strated in the urine and the amount of ammonia may vary from 8 to 1 2 gms. 
in 24 hours and represent from 25 to 40.4% of the total urine nitrogen; 
in periodic insanity, in which Edsall found it markedly reduced before the 
attack and increased just as the attack came on ; in certain severe cases of 
liver cirrhosis since the liver is no longer able to change it to urea ; in some 
cases of the pernicious vomiting of pregnancy in which the ammonia may 
represent even from 20 to 45% of the total urine nitrogen, while in cases 



THE URINE: THE NITROGENOUS BODIES 



123 



of nervous or reflex vomiting, and in eclampsia there may be no marked 
increase. (Definite hepatic lesions are found at autopsy in such cases.) 
And, finally, in normal pregnancy the ammonia percentage is somewhat 
increased and reaches its maximum during labor. 

Quantitative Determination of Ammonia. — The Schlosing method as 
modified by Schaffer is simple and fairly accurate, but too time consuming, 
(see Fig. 28). The apparatus consists of a wide crystallizing dish or wide 
Petri's dish, B, from 15 to 17 cm. in diameter so that the urine need not 
be over 2 mm. deep. This rests on a thick glass plate with accurately 
ground surface. Above the dish of urine, on a triangle, is another dish, 




Fig. 28. 



-Ammonia determination, Schlosing's method. A, bell- jar; B, dish containing urine; C, dish 
containing acid. 



C, into which have been previously measured 20 c.c. of o. iN H 2 SO 4 . These 
dishes are to be covered by a bell-jar the cavity of which is to be made air- 
tight by greasing its edge and the glass plate thoroughly. 

To 25 c.c. of filtered urine measured into the dish B, are added 0.5 gm. 
of sodium carbonate plus an excess of sodium chloride. The sodium carbon- 
ate will not split off ammonia from any of the other nitrogenous compounds, 
as for instance urea, and the sodium chloride will prevent decomposition. 

As soon as the alkali has been added the dishes are covered by the bell- 
jar and the apparatus then not disturbed for 3 or 4 days, or 48 hours if the 
apparatus be kept at 3 8° C, during which time the sodium carbonate will 
have set free all of the ammonia and the sulphuric acid will have taken 
it up. At the end of this time the sulphuric acid is titrated against 0.1N 



124 



CLINICAL DIAGNOSIS 



sodium hydroxide to determine the amount of acid which has been neutral- 
ized by the ammonia. This figure multiplied by 1.7 mgms. equals the 
weight of the ammonia originally in the 25 c.c. of urine. If any moisture is 
visible on the inside of the bell- jar the reaction of this should be tested with 
litmus and, if alkaline, the entire inner surface of the bell-jar should be 
washed with distilled water into the sulphuric acid before this is titrated. 

Folin's Method. — Folin's method is by far the best yet proposed for the 
determination of ammonia. 

Twenty-five cubic centimeters of urine are measured into an aerometer 
cylinder (see Fig. 29) from 30 to 45 cm. high and about a dram of dry 
sodium carbonate added. The further addition of from 5 to 10 c.c. of 
crude petroleum will prevent foaming. 



II 




B 






Fig. 29. 



-Folin's apparatus for ammonia and acetone determination. A, narrow tube for urine, con- 
nected by a tube containing cotton, with B, the cylinder containing acid. 



The upper end of this cylinder is then closed by a doubly perforated 
rubber stopper through which pass 2 glass tubes, only 1 of which is long 
enough to reach below the surface of the liquid. The shorter tube (about 
10 cm. in length) is connected with a glass tube which extends to the 
bottom of cylinder B (capacity about 500 c.c.) which contains 20 c.c. of 
0.1 N H2SO4, 200 c.c. of water, and 2 drops of a 1% solution of alizarin red 
as indicator. The special absorption device designed by Folin and pictured 
in Fig. 29 compels a very intimate contact of the air containing the ammonia 
with the acid through which it passes. The absorption bottle is now 
attached to a good filtering pump which can suck a strong current of air. 
The air passing through the alkaline urine and then through the standard 
acid will in about one and a half hours transfer every trace of ammonia 
to the acid. Its amount is then determined by direct titration with 0.1N 
NaOH, titrating to a red and not to a violet color. In titration involving 



THE URINE: THE NITROGENOUS BODIES 125 

ammonia phenolphthalein should not be used as indicator, but rather 
alizarin red, cochineal or a dilute solution of hematoxylin. 

Colorimetric Method. — The amount of urine to be used in this determin- 
ation should contain from 0.75 to 1.50 mgms. of ammonia nitrogen. While 
in the case of average normal urines 2 c.c. will be about the right amount, 
this will vary from 5 c.c. of very dilute to less than 1 c.c. of concentrated 
urines. The desired amount is measured with a pipet into a test-tube 
about 200 mm. in length and of such diameter that it will slip easily into 
a 100 c.c. narrow cylinder (B, Fig. 27). 

The apparatus used is that described on page 113 (see Fig. 27). Here 
also cylinder A contains 20 c.c. of distilled water and 2 to 3 drops of 10% 
hydrochloric acid. To the urine in the test-tube is added 1 c.c. of amy lie 
alcohol or 2 to 3 drops of caprylic alcohol (to prevent foaming) and then 
from 3 to 5 c.c. of saturated sodium carbonate solution which is to run 
gently down the tube under the urine so that none of the ammonia will 
escape. The test-tube is then quickly placed in cylinder B and the stopper 
quickly inserted with tube c reaching almost to the bottom of the test- 
tube E. After the apparatus is properly connected the suction through 
the apparatus is started slowly and the speed gradually increased so that 
at the end of about 5 minutes the air current is as rapid as the apparatus 
will stand. Aeration will be complete in from 15 to 20 minutes. The 
apparatus is then disconnected and cylinder A used for the final determina- 
tion, after any acid clinging to tube b has been washed back into the 
cylinder with 2 or 3 c.c. of distilled water. 

The standard solution is made up in a 50 c.c. volumetric flask as de- 
scribed on page 114. 

To the acid in cylinder A, which contains the ammonia of the urine, 
is added from 15 to 25 c.c. (depending upon the depth of color) of diluted 
Nessler's solution (1 to 5), and this diluted to 50 c.c, 100 c.c, etc., according 
to the depth of color. The colorimetric reading should be made at once. 
The calculation is made using Table I (page no) and the results recorded 
as in ammonia nitrogen. 

Example. — Suppose that 2 c.c. of urine were used, that the final dilution 
was to 100 c.c and that the reading was 69. This would indicate 0.70 mgm. 
of N in 2 c.c. of urine, or 0.35 mgm. of ammonia N in 1 c.c of urine 

Creatinin. — Creatinin, the aldehyde of creatin, is found in normal urine ; 
creatin seldom. In general its origin is the muscle ingested as food and 
the catabolism of body muscle. Its excretion runs roughly parallel to that 
of urea ; it is increased if the meat of the diet is increased and diminishes 
during fasting periods. Some claim that it is increased by excessive mus- 
cular work only, others (Edsall) that it is increased by all muscular exercise, 
and is diminished in diseases associated with extensive muscular paralysis, 
as well as by all conditions which markedly decrease the use of the muscles ; 
that while it is not a perfect index of the condition of muscle metabolism, 



126 CLINICAL DIAGNOSIS 

yet it is the best we have. Normally the output of creatinin in the urine 
is about i gm. a day. Folin 22 has shown that while the creatinin output 
of different normal persons on a meat-free diet varies widely yet that of 
each individual is an almost constant quantity which is quite independent 
of quantitative changes in the total amount of urine nitrogen. He found 
that moderately corpulent persons eliminated each 24 hours about 20 mgms. 
of creatinin and lean persons about 25 mgms. per kilo of- body weight. 
Folin also believes 23 that biologically at least creatin and creatinin are not 
related; that creatinin is a waste product, while creatin is treated as a food. 
Amberg and Morrill 24 found in the urine of the new-born a small but 
constant amount of creatinin. McCrudden 25 found in 3 cases of intestinal 
infantalism that the creatinin was more irregular than normal, that crea- 
tinin coefficient was low, and that creatin was present in the urine even 
when absent from the food. 

Jqffe's Test. — To the urine at room temperature is added a little aqueous 
solution of picric acid and a few drops of dilute NaOH. An intense red 
color at once develops which increases in intensity and then remains con- 
stant for hours. If acid be added the color becomes yellow. Acetone 
should previously have been removed by boiling. Glucose under these 
conditions gives, if warmed, a red color. The test is positive for creatinin 
in solutions of 1 to 5000 or more. 

WeyVs Test. — To the urine are added a few drops of very weak sodium 
nitroprusside (sp. gr. 1.003) an( i then a few drops of weak NaOH. A ruby- 
red color appears which soon changes to yellow. If acetic acid be next 
added and the fluid heated this color will change to green and then to a 
Berlin blue. This is positive for 0.6 gm. of creatinin per 1000 c.c. of urine. 
Acetone, if present, should be removed by boiling, for it would give a similar 
color, which, however, on the addition of acetic acid would change to a 
cherry-red or purple-red color. 

The most important compound of creatinin is its zinc salt (C4H7N 3 2 ) 2- 
ZnCl 2 . Creatinin on long boiling will decolorize Fehling's solution and 
after still longer boiling, if an excess of copper is present, will precipitate 
yellow Cu 2 (OH) 2 . It is to avoid this error when testing urine with Fehling's 
solution for glucose that we limit the time of boiling. Creatinin will not 
reduce a bismuth solution and this is one of the advantages of Nylander's 
test for glucose. 

Quantitative Determination of Creatinin — Folin's Quantitative 
Method.™ — This determination is based on the color reaction which 
creatinin (and no other normal urinary constituent) gives with picric acid 
in alkaline solution. Any fairly good colorimeter will suffice. 

22 Am. Jour, of Phys., 1905, vol. 13. 

23 Festschrift f. Olaf Hammarsten, iii, 1906. 

24 The Jour, of Biol. Chem., 1903, vol. iii, No. 4. 

25 Jour, of Exper. Med., Feb. 1, 1912, vol. xv, p. 107. 

26 Am. Jour, of Phys., 1905, vol. 13, p. 48. 



THE URINE: THE NITROGENOUS BODIES 127 

The reagents necessary are: a 0.3 N solution of potassium bichromate 
(which will contain 24.55 g ms - P er liter); a saturated picric acid solution 
(containing about 12 gms. per liter) and a 10% solution of sodium hydrate. 

Ten cubic centimeters of urine are measured into a 500 c.c. volumetric 
flask, 15 c.c. of the picric acid and 5 c.c. of the sodium hydrate solutions 
are then added, and the mixture is allowed to stand for 5 or 6 minutes. 

This interval is used to pour a little of the bichromate solution into 
each of the 2 cylinders of the colorimeter. The depth of the solution in 
1 of the cylinders is then accurately adjusted to the 8 mm. mark. With 
the solution in the other cylinder a few preliminary colorimetric readings 
are made simply for the sake of insuring greater accuracy in the subsequent 
readings of the unknown solution. No 2 readings should differ more than 
0.1 or 0.2 mm. from the correct value (8 mm. since both cylinders contain 
the same fluid) if we leave out of consideration the very first reading made, 
which is sometimes less accurate. Four or more readings should be made 
in each case and an average taken of all but the first. After a while 
one becomes sure of the true point, and can take the average of the 
first 2 readings. 

At the end of 5 minutes the contents in the 500 c.c. flask are diluted 
up to the 500 c.c. mark, the bichromate solution is thoroughly rinsed out 
of 1 of the cylinders by means of the unknown solution in the flask and 
several colorimetric readings then macle at once. 

The calculation of the result is very simple. It has been determined 
experimentally that 10 mgms. of perfectly pure creatinin will give under the 
conditions of the determination 500 c.c. of a solution of which a column 
8.1 mm. deep will have exactly the same colorimetric value as one 8 mm. 
deep of a 0.2N bichromate solution. If then for example it takes 9.5 mm. 
of the unknown urine-picrate solution to equal the 8 mm. of the bichromate, 

then the 10 c.c. of urine contain 10 X — = 8.4 mgms. of creatinin. If the 

9-5 
10 c.c. of urine examined are found to contain more than 15 mgms. or less 
than 5 mgms. of creatinin the determination should be repeated using corre- 
spondingly different amounts of urine in making up the 500 c.c. of fluid 
to be tested, since outside of these limits the results are much less accurate. 
This determination takes less than 15 minutes. 

Colorimetric Quantitative Determination of Creatinin. — Into a 100 c.c. 
volumetric flask or cylinder one measures with a pipet 2 c.c. of the urine 
to be examined. To this are added 3 c.c. of saturated picric acid and 1 c.c. 
of 10% sodium hydroxide, they are mixed thoroughly and allowed to stand 
for 5 minutes in order to allow the color to develop. At the end of this 
time the mixture is made up to 100 c.c. with tap water, thoroughly mixed 
and several readings made in the colorimeter, using as standard normal 
bichromate solution (made by dissolving 49.12 gms. of potassium bichro- 
mate in distilled water and making the solution up to 1 liter) . The amount 



128 



CLINICAL DIAGNOSIS 



of creatinin in 2 c.c. of urine is estimated by the use of Table III. If, e.g., 
the reading is 58, it indicates that the 2 c.c. of urine used and diluted to 
100 c.c. contain 1.62 ragms. and 1 c.c, 0.81 mgm. of creatinin. The creatinin- 
nitrogen would be 37.2% of this. If the concentration of creatinin in the 
urine is such that the readings do not fall within the figures of the table, the 
test is repeated using larger or smaller amounts of urine as the case may be. 

TABLE III * 



Estimation of Creatinin with the Hellige Colorimeter 



Colorimetric 
reading 


Creatinin 
mgms. per dilu- 
tion of 100 c.c. 


Colorimetric 
reading 


Creatinin 
mgms. per dilu- 
tion of 100 c.c. 


Colorimetric 
reading 


Creatinin 
mgms. per dilu- 
tion of 100 c.c. 


20 
21 
22 

23 
24 


2.46 

243 
2.4I 

2-39 

2-37 


35 
36 
37 
38 
39 


2.I3 
2.IO 
2.08 
206 
2.O4 


51 

52 

53 
54 
55 


I.78 
I.76 

1.74 
I.72 
I.69 


25 ' 

26 

27 

28 

29 


2-35 
2-33 
2.30 
2.28 
2.29 


40 
41 

42 
43 
44 


2.02 
1.99 
1.97 

i-95 
1.92 


56 
57 
58 
59 
60 


I.67 
1.65 
1.62 
I.60 

i-57 


30 
31 
32 

33 
34 


2.24 
2.21 
2.19 
2.17 
2.15 


45 
46 
48 
49 
50 


1.90 

1.88 
1.85 
1.83 
1.81 


61 
62 

63 
64 

65 


1-54 
1.51 

1.48 

1-45 

1.42 



* Myers and Fine's table copied from Gradwohl and Blaivas. 

Creatin. — For the quantitative determination of creatin, one measures 2 
c.c. of urine into a medium-sized test-tube and adds 2 c.c. of normal 
hydrochloric acid and a very little powdered metallic lead. The contents 
of the tube is now boiled nearly to dryness over a free flame and then 
washed with as little water as possible through a small cotton or glass-wool 
filter into a 100 c.c. volumetric flask. This removes the metallic lead which 
also reacts with the picric acid and alkali. All the creatin is now in the 
form of creatinin. To the fluid in the volumetric flask one now adds 3 c.c. 
of saturated picric acid and 2 c.c. of 10% sodium hydroxide, mixes thor- 
oughly and allows the solution to stand for 5 minutes. Then the mixture 
is made up to 100 c.c. with tap water, mixed thoroughly and read several 
times in the colorimeter, using the same standard solution (normal bichro- 
mate) and table as for creatinin. The result obtained is the total creatinin. 
The difference between the performed and the total creatinin gives the 
creatin in terms of creatinin, or, by multiplying this value by 1.16, the 
weight of the creatin. 

Oxyproteinic and Alloxyproteinic Acids. 27 — Oxyproteinic acid was 
isolated by Gottlieb and Bondzynski, and alloxyproteinic acid by Bond- 

27 Bondzynski and Panek, Ber. d. chem. Gesell., 1902, vol. xxxv, p. 2959. 



THE URINE: THE INORGANIC ACIDS AND BASES 129 

zynski and Panek. It is claimed that these acids contain all or nearly all 
of the neutral sulphur of the urine (they contain about 6% of sulphur) 
also that the oxyproteinic acid explains Ehrlich's Diazo reaction (see page 
157). These acids are said to stand the nearest to proteid of all the products 
of proteid metabolism and yet they give none of the proteid reactions. 
These writers claim that the normal urine contains about 1.2 gms. of the 
alloxyproteinic acid per day and about 3 times that amount of oxyproteinic 
acid. These acids have not been sufficiently studied as yet, and already 
some have been unable to confirm this work. 

THE INORGANIC ACIDS AND BASES 

The chlorides make up the bulk of the inorganic matter of the urine. 
In terms of the sodium salt the urine contains each 24 hours from 10 to 
15 gms. of sodium chloride. Practically all of the chlorine of the urine is 
present in inorganic combination, little if any in organic compound. 28 

The source of the chlorides of the urine is the food. Starvation will 
reduce them to a trace. More is excreted during the day than during the 
night. They are increased by anything which increases the amount of 
urine and also by active exercise. They are diminished during a period 
in which there is a loss of fluid to the body from diarrhea or vomiting 
and during the formation of a transudate or exudate and are increased 
while these are reabsorbing. In acute fevers the chlorides are usually 
diminished during the fastigium and increase during and after the defer- 
vescence. In acute lobar pneumonia there may not be a trace in the urine 
for several days before the crisis. The practically complete retention of 
chlorides has little or no prognostic value but has considerable for diag- 
nosis since in a case of doubtful fever it always suggests acute lobar pneu- 
monia. After the crisis the output soon returns to normal. The explana- 
tion of this phenomenon in pneumonia is not clear. It is not due to the 
diet, nor to lack of absorption from the bowel (for chlorides injected sub- 
cutaneously are retained) nor is all retained in the exudate. The chlorides 
in the blood are not increased but are in the other fluids of the body and 
are fixed in the tissues, even to 4 times the normal amount. 

With the febrile crisis of pneumonia the chlorides in the urine suddenly 
increase, even producing a " chlorine crisis," which may be the first sign 
of improvement. The sulphates and the phosphates may be retained to a 
lesser degree, but do not return to normal at the same time as do 
the chlorides. 

We have studied the records of 34 cases of acute lobar pneumonia in the Johns 
Hopkins Hospital, the total chloride output of whom was determined daily. Of these, 
1 1 were on a pure milk diet, 1500 c.c. per day. Six terminated by crisis. In 2 the chlor- 
ides showed a drop toward the crisis, in 1 the crisis was preceded by a rise, while in other 
cases the rise began with, or even 4 or 5 days later than, the fall in temperature. It will 

28 Ville and Moitessier Compt. rend. Soc de Biol., liii, p. 673. 
9 



130 CLINICAL DIAGNOSIS 

be seen that in these very few cases we obtained very little evidence of prognostic value 
from the determination of the chlorides. In no case were the chlorides entirely absent. 
On the day before the crisis they varied from 0.7 to 2.1 gms., average 1.3 gms. The 
greatest rise began on the fifth day after the crisis, on which day it varied from 3.8 
to 4.9 gms. 

Of 22 cases terminating by lysis the daily output of chlorides in J{ of the cases fell 
toward the lysis and in %> it began to rise 1 or 2 days before the temperature began to 
fall. On the first day of the lysis the output was above 1 gm. in 10 cases (averaging 
26 gms.) and 9 gms. in 1 case. The chlorides were entirely absent in 3 cases before 
defervescence and in 2 cases during the fall of the temperature. In these cases therefore 
their entire absence was not a bad sign. They reached their lowest point during the 
fall of temperature in )i of the cases, just before the beginning of the lysis in %, and began 
to rise with the lysis in just }i of the cases. The chief rise began and was most rapid 
after the temperature had reached normal. 

In 5 fatal cases the chlorides fell steadily until death in 3 cases, rose in 1, while in 
1 case death was preceded by 6 days during which they were quite absent. 

In 1 case of delayed resolution the chloride curve was very interesting. Nineteen 
determinations were made during a period of twenty-two days. The lowest amount 
was 4.3 gms. and this was after the lysis. For the most part it varied from about 5 to 
10 gms. per day. In this case there was therefore comparatively little chloride retention. 

In those cases in which the lysis or crisis is followed by several days of very slight 
fever the chlorides may not rise until the temperature is quite normal. In other cases 
with a normal temperature but with a continuous slight leucocytosis they did not rise 
until this had fallen below 10,000. Ev? 

The chlorides of the urine are increased after chloroform inhalation, 
in diabetes insipidus and in other conditions with marked polyuria. They 
are decreased in all chronic diseases, in which cases the reason may be dis- 
turbed absorption, the diet, or the condition "of the kidneys. They are 
decreased also in gastric diseases when there is considerable vomiting, when 
absorption is diminished as in malignant pyloric stricture, and when they 
are lost to the body by lavage or diarrhea. 

It is an ominous sign in chronic diseases if the output of chlorides drops 
to as low as 2 gms. (provided the diet cannot explain this) for the cessation 
of chloride elimination sometimes is a sign of on-coming death. 

The output is very low in meningitis and only moderately low in typhoid 
fever. It is markedly diminished in cholera, pyemia, puerperal fever, in 
serum disease 29 and in acute articular rheumatism. In cirrhosis of the 
liver it is said to be increased. The explanation by Widal of edema in 
nephritis as due to a specific retention of chlorides because of renal insuffi- 
ciency, while the output of other solids remains normal, was accepted for 
several years and then abandoned. Those who adopted this theory applied 
the term " chloruremia " to a partial renal insufficiency for chlorine 
elimination, with a rapidly developing general edema, a low CI output 
and an increasing output of albumin in the urine. It would be hard to 
explain on this basis the absence of edema after even a week of total suppres- 
sion of the urine, e.g., to calculus, or to the removal by operation of the 

29 Rackemann, Longcope and Peters, Arch, of Int. Med.; Oct., 1916, vol. xviii, p. 496. 



THE URINE: THE INORGANIC ACIDS AND BASES 131 

only functioning kidney. We have repeated the work of Widal with varying 
success, but with no success at all if the water intake was controlled and 
this was difficult since the salt makes the patient very thirsty. 

Estimation. — A rough estimation of the amount of chlorides in a 
specimen of urine is easily made by dropping i drop of AgN0 3 solution 
(1:18) into a test-tube of clear urine which contains no albumin and to 
which 10 drops of pure nitric acid had been added. If the chlorides are 
normal or increased in amount, the precipitate forms as a compact ball 
which sinks to the bottom; if diminished, this ball is less compact; if much 
diminished, e.g., to 0.1% or less, only a cloud will be produced. 

Quantitative Determination. — The best method of determining the 
chlorides of the urine is Arnold's modification of Volhardt's method. With 
the chlorides are estimated also the minute traces of cyanides present. The 
urine should contain no nitrites, and most observers add also, no albumin or 
albumose, since these will be precipitated as silver albuminates. If albumin 
be present, it may be necessary to ash the urine (Neubauer's method). 
Hammarsten recommends that the albumin be removed by heat and 
acetic acid, but the albumin precipitate must be washed for some time in 
order that the abundant chlorides which it will contain may be regained. 

This method is as follows : The chlorides are precipitated by an excess 
of silver nitrate in a urine made strongly acid by nitric acid. The precipi- 
tate is then filtered out and the excess of silver nitrate determined by 
titration with ammonium sulphocyanate. 

vSolutions necessary: (i) An AgN0 3 solution i c.c. of which will exactly 
precipitate 10 mgms. of NaCl. The pure crystallized AgN0 3 is used, 
29.075 gms. dissolved in 1 liter of distilled water. 

(2 ) A cold sataated solution of iron ammonium alum, or ferric sulphate, 
chlorine free (50 gms. of Fe 2 6 per liter). 

(3) HNO3, specific gravity 1.2, chlorine-free. If chlorine be present 
the acid should be purified by distillation. Any nitrous acid should be 
removed by the addition of urea. 

(4) An ammonium sulphocyanate solution 10 c.c. of which will equal 
10 c.c. of the silver nitrate solution. To obtain this, 12.9 gms. of the 
NH 4 SCN are weighed and dissolved in a little less than 1 liter of water. 
Twenty cubic centimeters of the silver nitrate solution, 5 c.c. of the iron 
alum, and 4 c.c. of nitric acid are mixed in a flask and then diluted to 100 c.c. 
The ammonium sulphocyanate solution is then added from a buret. The 
precipitate, at first brown, at once changes to the white precipitate of silver 
cyanate, until the last particle of silver has been precipitated after which 
the brown ferric cyanate precipitate will persist. This end reaction is 
very sharp. The volume of the solution may then be corrected. Others 
recommend (v. Jaksch) that this solution be so made up that 25 c.c. of 
it will equal 10 c.c. of the silver nitrate, while others, that 20 c.c. equal 
10 of the silver solution. 



132 CLINICAL DIAGNOSIS 

To determine its chlorides 10 c.c. of the urine are carefully measured 
with a pipet into a ioo c.c. measuring flask. Then are added 20 to 30 
drops of nitric acid and 2 c.c. of the iron alum solution. If highly colored 
a few drops of 8% KMn0 4 also are added. The silver nitrate solution is 
then slowly run in from a buret, constantly shaking the flask until one is 
sure that all the chlorine has been precipitated and that there is an excess 
of the silver solution. It is usually safe to add 20 c.c. of the silver solution, 
while others recommend that 15 c.c. be used. In general the greater the 
excess the better the results. The flask is then allowed to stand for about 
10 minutes, then filled to the 100 c.c. mark with water and thoroughly 
mixed. There should be an excess of iron present otherwise the nitric add 
will decolorize the ferric cyanate but this excess of iron causes a brown 
rather than a red color in the end reaction. 

The contents of the flask is then filtered through a dry filter until 
50 c.c. of clear filtrate are obtained. This is titrated with the ammonium 
sulphocyanate solution until the end reaction. The amount used indicates 
the excess of the silver solution in 50 c.c. of filtrate. This amount multi- 
plied by 2, since only % of the filtrate was used, and subtracted from the 
number of cubic centimeters of silver nitrate originally added, will give the 
number of cubic centimeters of silver nitrate actually precipitated by the 
chlorides of the urine. This multiplied by 10 mg. will give the weight of 
the chlorides as sodium chloride in the amount of urine used. 

Some add the iron-alum solution to the 50 c.c. of filtrate, not before. 
A much- jaundiced urine should be decolorized by adding a few drops of 
potassium permanganate and nitric acid. The urine is then warmed, 
allowed to stand for a few minutes, and filtered. 

Harvey's Method.— Harvey 30 recommends a modification of the Volhard 
method which seems accurate, and which certainly is much simpler. 

The solutions used are the silver solution described above, a sulpho- 
cyanate solution prepared as above, 20 c.c. of which are equivalent to 
10 c.c. of the solution of silver nitrate, and a third solution, the " acidified 
indicator," which is prepared as follows: To 30 c.c. of water are added 
70 c.c. of nitric acid (sp. gr. 1.2, or 33%). One hundred grams of crystalline 
ferric ammonium sulphate are dissolved in this menstruum, and the solution 
is then filtered. 

Five cubic centimeters of the urine are pipeted into a small beaker 
and diluted with about 20 c.c. of distilled water. The chlorides are now 
precipitated with exactly 10 c.c. of the solution of silver nitrate, and about 
2 c.c. of the acidified indicator are added. The solution of ammonium 
sulphocyanate is then run in from a buret until the first. trace of red per- 
sists throughout the mixture. The number of cubic centimeters of the 
sulphocyanate solution used is divided by two (since 20 c.c. of this solution 
equal 10 c.c. of the silver solution), and this quotient subtracted from 10. 

30 Arch, of Int. Med., July 15, 1910, vol. 6, p. 12. 



THE URINE: THE INORGANIC ACIDS AND BASES 133 

The difference is the amount of the silver nitrate solution used to precipitate 
the chlorides in the urine. Each c.c. of this is equivalent to o.oi gm. of NaCl. 

Phosphates. — The amount of phosphates weighed as P2O5 in the urine 
of an adult varies from 1 to 5 gms., average about 3.5 gms., in 24 hours. 
Of this the earthy phosphates are estimated as from 1 to 1.5 gms. and the 
alkaline from 2 to 4 gms. These form a constant sediment in an alkaline 
urine, a constituent of some of the most common crystals and the principal 
ingredient of some of the commonest stones. In addition to the mineral 
phosphates there is always in the urine a little phosphorus in organic 
combination. This amount in the urine depends especially on the amount 
in food and also inversely as the food's content of calcium and magnesium 
which form in the intestines insoluble phosphates, the most of which are 
eliminated with the stools. (Much of the acid calcium phosphate is ab- 
sorbed from the bowel.) Because of this the phosphates of the urine may 
decrease to even less than 1 gm. in 24 hours. This is the reason why the 
urine of certain of the herbivora contains only traces of phosphoric acid. 
In metabolism experiments involving phosphoric acid one should control 
the diet carefully that the amount of this acid absorbed be approxi- 
mately constant. 

The phosphate output is increased by a nuclein-rich diet; by any con- 
dition which increases the metabolism of the body tissues (the amount 
from this source, however, is small) ; and by hard muscular work. In 
starvation it falls somewhat, yet less than the nitrogen. In dogs on a 
pure meat diet, N : P2O5 : : 8.1 : 1. 

Clinically the phosphates of the urine have been the subject of much 
discussion. Some state that they are increased in extensive disease of bones, 
as rickets, osteomalacia, diffuse periostitis, etc.; in destructive disease of 
the lungs, especially in early tuberculosis; and in extensive disease of the 
nervous system. Nevertheless the evidence is not clear as regards any 
one of these diseases. In mental disease Folin and Shafer 31 found that 
while during the periods of excitement the output of phosphoric acid might 
be relatively diminished, yet there was but little absolute change. 

It would seem to be increased in meningitis, in yellow atrophy of the 
liver, in diabetes mellitus, in diabetes insipidus, after the use of chloral, 
of KBr, and lastly in phosphorus poisoning. 

It has been found diminished; in pneumonia during the fastigium (in 
children, however, there may be a rise (v. Jaksch)) and increased with the 
crisis (but not always simultaneous with the rise of nitrogen and of chlorine ; 
at this time the ratio between the earthy and the total phosphates may 
increase considerably. Since in tuberculosis these are said to be increased, 
Gouraud suggests this as of aid in the differential diagnosis between these 
two conditions) ; in typhoid fever (in 1 case the total P2O5 rose after defer- 
vescence from 1.5 to 13 gms.) ; in most chronic diseases, especially in renal 
31 Am. Jour, of Physiol., vol. vii, p. 135. 



134 CLINICAL DIAGNOSIS 

disease (Purdy stated that in nephritis a diminution in the output of 
phosphate is almost as constant as is the albuminuria) ; in pregnancy, in 
which case it is attributed to the fetal bone formation; and in gout, in which 
disease the line of phosphoric acid elimination runs quite parallel to that 
of uric acid. And yet in all these diseases the output of phosphates is 
subject to large and sudden variations which are independent of the diet. 
Certain cases have been reported (Teissier) with all the symptoms of 
diabetes mellitus, except glycosuria, and a phosphate excretion reaching 
even 10 gms. in 24 hours, the so-called " phosphatic diabetes." This is a 
term applied by some to cases with a minimal output of the phosphates 
of at least from 3.5 to 4 gms. per day, but by others to cases in which 
P 2 5 : N : : 17-20 : 100. Some say that these cases are merely neuras- 
thenic ; others that they are definitely diabetic with a rich phosphate output 
during the sugar-free periods. 32 

The phosphorus which appears in the urine in organic combinations 
is apparently not influenced by a phosphorus rich diet, but would seem to 
be a good index of tissue catabolism (Mandel and Oertal) . 

In clinical chemistry one meets with 4 groups of phosphates — the 
diacid, monacid, normal and basic. These vary in solubility in the order 
in which they are named, the diacid being the most so. The monacid 
phosphates of calcium and magnesia explain the cloud which appears 
when urine is made alkaline by the addition of an alkali. The flocculent 
precipitate which appears when urine is heated and which resembles the 
cloud of albumin but is soluble in acetic acid, consists of normal calcium 
phosphate (basic, says v. Jaksch) with a trace of CaOx and CaSCU, but no 
magnesium salts since its salts are more soluble than are those of calcium. 

In leukemia, White and Hopkins 33 found the phosphate diminished 
both absolutely and relatively (to nitrogen) and suggest that the phos- 
phorus is retained in the body to build new leucocytes. In the new-born 
the proportion between nitrogen and phosphoric acid is from 5 to 8 : 1 . 

Of the normal phosphates that of greatest interest is MgNH 4 P0 4 6H 2 
of which the beautiful coffin-lid crystals of triple phosphate are composed 
(see page 248). 

The acidity of the urine, due to many acid bodies and in an unknown 
degree to each, is, however, due chiefly to acid phosphates. Normally 
60% of the phosphoric acid of perfectly fresh urine is present as diacid- 
phosphate and 40% as the monacid salts, but the former varies from 34.9 
to 74.2%. In general it may be said that the urine will be amphoteric if 
the diacid salts are from 30 to 50% and the monacid from 70 to 50% of 
the total phosphate output. 

An easy approximate quantitative determination of the phosphates of the 
urine is made by filling a test-tube half full of filtered urine, adding am- 

32 Ralfe, Lancet, Mar. 5, 1887. 

33 Journal of Physiology, vol. xxiv, p. 42. 






THE URINE: THE INORGANIC ACIDS AND BASES 135 

monia, warming and then allowing it to stand. If in from 18 to 24 hours 
the precipitate of the earthy phosphates is from % to % an inch deep, the 
amount is normal ; if less, it is diminished. This is then filtered out, all of 
the filtrate put back in this test-tube and 1 finger's breadth of magnesium 
mixture added. The urine is then warmed and the precipitate of alkaline 
phosphates allowed to settle. If during the same length of time the sedi- 
ment is from % to % inches deep the amount is normal. 

A urine may -be cleared of phosphates by precipitating it with basic or 
neutral lead acetate. 

Quantitative Determination; Uranium Nitrate Method. — Phos- 
phoric acid is precipitated as a diacid salt by uranium nitrate. If cochineal 
is used as indicator the first excess of the uranium salt over and above that 
necessary to precipitate the phosphoric acid will give with the cochineal 
a green compound which serves as the end reaction. The uranium nitrate 
solution, although more stable than the acetate, should be frequently 
restandardized. Since the free nitric acid liberated in the reaction would 
redissolve a certain amount of uranium phosphate sodium acetate is added 
in excess ; and in order that all phosphoric acid may be present as a diacid 
salt, acetic acid is added as well. The urine should be titrated while boiling 
since the end reaction will be quicker and sharper. 

Albumin and sugar if present need not be removed. This titer changed 
with the volume of reagent used. For instance, if 20 c.c. are used, 1 c.c. 
will indicate 4.98 mgms. of P2O5; 21 c.c, 5 mgms.; 40 c.c, 5.14 mgms., 
etc For this reason the uranium nitrate solution should be standardized 
against a phosphoric acid solution which has about the same concen- 
tration as normal urine. 

The fluids necessary are: 1. A phosphate solution 50 c.c of which 
contain 0.1 gm. of P2O5. This is so difficult to prepare that we recommend 
that it be purchased from those chemists who make a specialty of such 
work. This is the standard solution. 

2. A solution containing 100 gms. of NaAc and 30 gms. of acetic acid, 
in 1 liter of water. Five cubic centimeters of this fluid added to 50 c.c 
of urine will keep all the phosphates in the diacid condition and prevent 
the presence during the titration of any free nitric acid. 

3 . An alcohol-cochineal extract made by digesting the pulverized insects 
in 25% alcohol. 

4. Uranium nitrate solution, 1 liter of which contains 35.461 gms. of 
U0 2 (NO 3)26^0. This solution is so standardized against solution 1 
that 20 c.c. of this solution will equal exactly 50 c.c of solution 1, that is, 
will indicate 0.1 gm. of P 2 5 . Three grams of NaAc are added since this 
salt always contains some free nitric acid. 

To 50 c.c of solution 1 in an Erlenmeyer flask are added 5 c.c. of solu- 
tion 2, then a few drops too much rather than too little of the cochineal 
tincture. This fluid is then kept at the boiling point on a water bath or 



136 CLINICAL DIAGNOSIS 

over a free flame, while the uranium solution is added from a buret in 
small amounts, shaking constantly. When near the end the precipitate 
which falls after each addition is allowed to settle somewhat and the bottom 
of the flask studied for the first trace of a green precipitate. Having deter- 
mined how much of this solution will exactly precipitate the phosphoric 
acid in 50 c.c. of solution No. 1, the proper correction is then made and the 
result again verified that the result may be 20 c.c. 

For the estimation of phosphoric acid in the urine 50 c.c. of this are 
treated in exactly the above manner. 

If very accurate results are desired a table of corrections for the change 
in titer necessary for the volume used should be used. 

If the urine is jaundiced or so colored that the end reaction is not sharp, 
it should be acidified with hydrochloric or nitric acid, decolorized with 
KMnO 4 and again neutralized. 

Sulphates. — Sulphur is present in the urine in 3 forms — (a) preformed 
or neutral sulphates; (6) ethereal or conjugated sulphate, that is, sulphuric 
acid combined with aromatic alcohols, indoxyl, skatoxyl, cresol, phenol, 
etc. ; (c) unoxidized, or organic sulphur. The normal person on a mixed 
diet eliminates from 1.5 to 3 gms., an average of 2.5 gms., of " a " and " b " 
(weighed as SO 3) in 24 hours. As a rule the ethereal sulphates make up 
about one-tenth of the total sulphates. Practically all of the sulphuric 
acid of the urine is a product of proteid metabolism, therefore its output 
runs parallel to that of nitrogen, and the ratio between them is quite, but 
not exactly, constant (100 : 19. 1-20.4 Folin). 

The total sulphate output is increased : in all conditions with increasing 
proteid oxidization, as a diet rich in meat; after exercise, providing this 
increases the nitrogen output as well; in fevers, since in this condition there 
is an increased proteid catabolism (especially in acute mflarnmatory disease 
of the brain and cord and in acute articular rheumatism) ; and after the 
ingestion of protoplasmic poisons. It is diminished during convalescence 
from an acute fever and in practically all chronic diseases. The amount 
of total sulphates has very little clinical value. 

The ethereal sulphates have attracted too much interest. While 
their output is subject to great and inexplicable variation, on the whole 
they are a fairly accurate index of the absorption of those products of intesti- 
nal decomposition which can pair with this acid. Among these are phenol, 
cresol, indoxyl, hydrochinin and pyrocatechin. While the most important 
of these are indoxyl and phenol yet these explain but about one-fifth 
of all the ethereal sulphate in the urine. Those which pair with the larger 
percentage of the sulphuric acid are still unidentified. It is the absolute 
not the relative amount of ethereal sulphates in the urine which is of value. 
Their output varies with the food. It is greatly increased by bad food 
and is diminished during periods of fasting. It is decreased by milk diet, 
since casein inhibits the growth of the bacteria of decomposition. Intestinal 



THE URINE: THE INORGANIC ACIDS AND BASES 137 

decomposition increases them both relatively and absolutely while calomel 
and similar drugs will at once decrease them. They are increased by the 
ingestion of aromatic bodies, especially of carbolic acid. There is almost 
none in the urine of the new-born. Their output is low when there is 
abundant hydrochloric acid in the gastric juice and is diminished by 
hydrochloric acid medication but increased by the ingestion of alkalies. 

Pathologically, the ethereal sulphates are increased: in chronic colitis 
(and diminished in the acute), in constipation often but not always (in 
an interesting specimen of " black urine " from a case of extreme constipa- 
tion the total sulphuric acid (as S0 3 ) was only 0.147 gm. per 100 c.c. of 
urine and of this 57% was ethereal sulphate; the following day the urine 
was of normal color and the total SO 3 was 0.086 gm. per 100 c.c. 50% of 
which was ethereal sulphate); in typhoid fever; intestinal tuberculosis; 
peritonitis ; cholera (but during the stage of reaction little or none may be 
present) ; in atrophic liver cirrhosis and in carcinoma of the liver (in which 
cases the increase is attributed to the accompanying intestinal disturbance) ; 
and as a result of decomposition in other parts of the body than the intestine. 
It is of interest that in gastric disease, even in cases with much stagnation 
and fermentation, they are little affected. 

The amount of unoxidized (e.g., organic or neutral) sulphur in 
the urine is supposed by some to vary with the amount and quality of 
the food ; by others, to have relation not to food but to body tissue 
catabolism ; to be increased by muscular work, by conditions leading to the 
oxygen starvation of the tissues and by the ingestion of various sulphur 
compounds, including the flower of sulphur, sulphonal, methylmerkaptan 
and ethyl sulphide. 

The organic sulphur, which amounts to from 14 to 25% of the total 
sulphur, is present in an easily oxidizable fraction, which is oxidized by 
bromine or chlorine (bromine is better, since chlorine attacks also the 
taurin derivatives), and a difficultly oxidizable fraction. 

To determine the total organic sulphur the dried residue after the 
removal of the sulphates must be fused with KN0 3 . Fuming HN0 3 will 
not oxidize all of it. In cystinuria HCI+KNO3 will oxidize only from 
30 to 40% of it. One then determines it as sulphuric acid. 

In jaundice from 24 to 60% of the sulphur in the urine is in unoxidized 
form and of this the difficultly oxidizable form is increased to about 4 to 5 
times its normal proportion. In pneumonia it is increased and in liver 
disease, decreased. In cystinuria even 45.7% (in 1 of our cases 32 %) of the 
sulphur in the urine is in neutral form. 

Edsall 34 who carefully studied the easily split (by alkali) sulphur in a 
series of cases decided that cystinuria is the only disease with an increase 
in the sulphur fraction and that the relative proportion of these 2 fractions 
has no clinical value. 

34 Univ. of Penn. Med. Bull., 1892, iii, p. 87. 



138 CLINICAL DIAGNOSIS 

Recent work by several (e.g., Benedikt) 35 has emphasized the independ- 
ence between the neutral and total sulphur and the possible origin of the 
former in the catabolism of particular proteids. 

Detection and Approximate Estimation. — In a test-tube of over 
25 c.c. capacity is mixed the urine and about one-third its volume of an 
acid barium chloride solution (BaCl 2 , 4; HCl,i; H 2 0, 16 parts). If the 
precipitate of neutral barium sulphate gives the urine a milky turbidity, 
the sulphates are normal; if creamy, increased; if merely a translucency, 
they are diminished. If the precipitate after settling from 18 to 24 hours 
fills % of the concavity of the tube the sulphates are normal. If this pre- 
cipitate be removed by filtration, hydrochloric acid added to the filtrate 
and the fluid warmed the ethereal sulphates are split and precipitated as 
neutral sulphates. 

Quantitative Determinations of Sulphur-containing Bodies. — The deter- 
minations of the sulphur bodies of the urine, the neutral, the ethereal sulphates and the 
total sulphur, are so important in metabolism experiments that the student should be 
trained in the general methods of this work. He should remember that these determina- 
tions, theoretically so easy and accurate, are in reality very difficult and so full of pitfalls 
that much preliminary practice should precede any important work. We have copied 
almost in full the method of Folin. 36 

Inorganic Sulphates. — About 100 c.c. of water (not less) 10 c.c. of dilute hydro- 
chloric acid (1 part of concentrated HC1 to 4 parts H 2 by volume) and 25 c.c. of urine 
are measured into an Erlenmeyer flask (capacity 200-250 c.c). If the urine is dilute, 
50 c.c. instead of 25, and a correspondingly smaller quantity of water may be taken. 
A 5% solution of barium chloride solution (10 c.c.) is then added, always drop by drop, 
preferably by means of an automatic dropper. The urine solution is not to be shaken, 
stirred or otherwise disturbed while the barium chloride is being added. At the end of 
an hour or later, according to convenience, the mixture is shaken up and filtered through 
a Gooch crucible. The precipitate is washed with about 250 c.c. of cold water, dried 
and ignited. Folin used porcelain crucibles rather than platinum and described the 
following technic: The asbestos for the mats must be good material, consisting chiefly 
of long shiny fibers. The fibers are cut with scissors into suitable lengths (50-70 mm.). 
A few grams at a time are then placed in a cylinder with about 300 c.c. of 5% hydro- 
chloric acid and a strong air current is passed through for a few minutes. This separates 
all the fibers far more quickly and completely than the usual method of scraping them 
with a knife. In an hour or 2 asbestos enough for 200 crucibles can be prepared. It is 
kept ready for use in dilute hydrochloric acid. From 50 to 100 mgs. of asbestos are 
used for each mat. By using a good vacuum pump at almost full force the asbestos 
mat is packed into a thin but uniform and firm layer in the bottom of the crucible. It 
is then washed with the help of only enough of a vacuum to make the water run through 
in a slow stream; dried, ignited, and weighed. Mats so prepared are as effective as the 
best filter paper in retaining precipitates and there is practically no danger of losing 
any asbestos during the subsequent washings of precipitates of barium sulphate. The 
same mat can be used until about 1 gm. of barium sulphate has collected. Time is saved 
by not using the same mat too long because the filtration becomes slower and slower 
the more precipitate there is present and it is not safe to increase the vacuum too much. 
The ignition of the precipitates is associated with more serious sources of error 
than the filtration, more serious because they are not accessible to direct observation. 

35 Zeitsch. f. Id. Med., 1899, vol. xxxvi, p. 281. 

36 The Jour, of Biol. Chem., Jan. 1906, vol. i, p. 131. 



THE URINE: THE INORGANIC ACIDS AND BASES 139 

The flame must not be applied directly to the perforated bottom of the crucibles. If 
this is done mechanical losses are sure to occur, even though the crucibles are covered 
with lids. Nor is it safe to apply the flame to sides of the crucibles. To do so involves 
again mechanical loss of barium sulphate. During the ignition the crucibles must be 
provided not only with lids, but also with tight bottoms. This is easily accomplished 
by the use of lids of ordinary platinum crucibles. The lid is placed on a triangle and the 
crucible stands in upright position on top, while the flame is applied to the platinum lid. 
Ten minutes ignition is sufficient unless organic matter is present. 

Total Sulphates (Neutral, i.e., Inorganic and Ethereal). — Of the following two 
methods Folin prefers the first : a. Barium Sulphate Precipitation in the Cold. — Twenty- 
five cubic centimeters of urine and 20 c.c. of dilute HC1 (1 part HC1, sp. gr. 1.20, to 
4 parts H 2 0), or 50 c.c. of urine and 4 c.c. of concentrated hydrochloric acid are gently 
boiled in an Erlenmeyer flask (capacity 200-250 c.c.) for 20 to 30 minutes (not less than 
20). To reduce the loss of steam it is better to keep the flask covered with a small watch 
glass during the boiling. The flask is cooled for 2 or 3 minutes in running water and the 
contents are diluted with cold water to about 150 c.c. To this solution is then added 
5% barium chloride (10 c.c.) without any shaking or stirring during the addition. The 
remainder of the procedure is like that of the inorganic sulphate determination. 

b. Barium Sulphate Precipitation in the Heat. — The boiling of the urine with hydro- 
chloric acid is conducted exactly as in the preceding method. At the end of 20 to 30 
minutes the boiling urine is diluted to about 150 c.c. with hot water. The mixture is 
heated once more to the boiling point, is then taken off the fire and at once precipitated 
with 10% barium chloride solution (5 c.c). The barium chloride must always be added 
drop by drop. The filtration is made after about 2 hours' standing, when the mixture 
has acquired the room temperature. The remainder of the procedure is like that of the 
determination of inorganic sulphates. 

Ethereal Sulphates. — There is no need that these be separately determined 
since the difference between the amounts of total and neutral sulphates found in any 
given specimen will be an accurate index of the ethereal sulphate in that partic- 
ular specimen. 

Total Sulphur. — The determination of the total sulphur is one of the most im- 
portant but most difficult problems of proteid metabolism. Folin's method is the following : 

Twenty-five cubic centimeters of urine (or 50 c.c. if very dilute) are measured into 
a large nickel crucible (capacity 200-250 c.c.) and about 2 gms. of sodium peroxide are 
added. The mixture is evaporated to a syrupy consistency and is then carefully heated 
until it solidifies. This heating may seem a little slow, requiring about 15 minutes, 
but the conditions have purposely been selected to make it slow (by using as much as 
3 gms. of Na20 2 ) in order to drive off as much ammonia as possible before the final 
fusion with more peroxide. The crucible is removed from the flame and allowed to cool. 
The residue is then moistened with 1 or 2 c.c. of water and, after about 7 gms. of sodium 
peroxide are sprinkled over the contents in the crucible, the mixture is heated to complete 
fusion for about 10 minutes. After cooling for a few minutes water is added to the con- 
tents in the crucible and the mixture is heated for at least half an hour with about 100 c.c. 
of water to dissolve the alkali and to decompose the sodium peroxide. The mixture is 
next rinsed into an Erlenmeyer flask (capacity 400-450 c.c.) by means of hot water 
and diluted to about 250 c.c. Concentrated hydrochloric acid is slowly added to 
the almost boiling solution until the nickelic oxide just dissolves (about 18 c.c. of acid 
to 8 gms. of peroxide). After a few minutes' boiling the solution should be perfectly 
clear. If it is not clear too much water or too little peroxide has been added for the 
final fusion. The insoluble residue must then be removed by filtration (after cooling) 
because it will not dissolve on the addition of more hydrochloric acid and too much 
acid must be avoided. The difficulty does not arise if little water and 7 or 8 gms. of 
peroxide are used. 



140 CLINICAL DIAGNOSIS 

To the clear acid solution are added 5 c.c. of very dilute alcohol (1 part alcohol to 
4 parts H2O) and the boiling is continued for a few minutes. The alcohol removes the 
last traces of chlorine which is always freed on acidifying the solution. Ten per cent, 
barium chloride solution (10 c.c.) is next added (by means of a dropper) and the solution 
left standing in the cold for 2 days before filtering. The rest of the procedure is the same 
as for the other sulphate determinations. 

Thiosulphuric Acid, H 2 S 2 3 . — Normally there is either no thiosulphuric 
acid in the urine or not over 10 mg. in 1 liter. It has been found in some 
cases, as in typhoid fever. 

Hydrogen Sulphide, H 2 S. — This gas is seldom present in fresh urine. 
It does, however, occur and to it have been attributed cases of autointoxi- 
cation. It was found in 1 case of long-standing eclamptic coma. It soon 
appears, however, in almost any urine on standing and may be detected 
by its odor, or by suspending in the mouth of the flask containing the 
urine a strip of paper moistened with sugar of lead solution plus 1 drop 
of NaOH. Air should then be blown through the urine. The paper will 
be blackened. 

Sulphocyanic Acid, HSCN. — This acid occurs normally in the urine of 
man and of the animals which excrete nitrogen as urea, in amounts equalling 
about one-third that of the neutral sulphur. To demonstrate it, 100 c.c. 
of urine are precipitated by HNO3, filtered, the precipitate washed, 
suspended in water, decomposed with H 2 S and this nitrate distilled. 
The distillate, tested with Fe 2 Cl 6 , gives an intense blue color (Berlin blue) 
not modified by HC1. 

Carbonates. — Carbonic acid is present in the urine, free, i.e., which may 
be removed by a vacuum, and bound, in which case acid must be added to 
free it. Of the free, the urine contains about 180 c.c; of the bound, from 
2 to 10 c.c. The carbonic acid is increased by a diet rich in those organic 
acids which are oxidized to carbonates, hence there is much in the urine 
of the herbivora. 

Silicic acid is present in traces as silicates. Its source is the food. 
Nitric acid is present in all normal urines as nitrates. This is from the food. 
Nitrous acid is often found in the urine as nitrates, but this was eliminated as nitrates 
and later reduced by bacteria. 

Calcium and Magnesium. — The phosphates of the alkaline earths, 
calcium and magnesium, are present in the urine to the amount of about 
1 gm. per day; calcium, weighed as CaO, about 0.12 to 0.25 gm. ; and MgO 
from 0.18 to 0.28 gm. per day. Calcium, even that injected subcutaneously, 
is excreted chiefly through the intestinal wall and only about 4 to 29% 
through the kidneys, therefore the output in the urine is no index of the 
amount ingested which was used. Under normal conditions the most of 
the calcium eliminated is from the food. There is only a trace in the urine 
of persons on a vegetable diet. Its output in the urine runs parallel to that 
of ammonia and seems directly related to the excretion of acids. It seems 






THE URINE: THE INORGANIC ACIDS AND BASES 141 

to be increased by exercise. The most of that in the urine is eliminated in 
the morning, at which time the urine is most acid. It is increased both 
relatively and absolutely during periods of starvation when a slight acidosis 
is always present. This calcium is supposed to come from the bones. It 
can be decreased by the administration of alkalies. 

The factors influencing the output of calcium in disease are but little 
understood. There is no increase in tuberculosis and none in rickets. The 
contradictory findings in the chronic diseases which have been the subject 
of so much careful study can best be attributed to the inanition which 
these diseases cause. In diabetes, and especially in the cases with acidosis, 
Gerhardt and Schlesinger found that the output in the urine is increased to 
even 2 to 4 times the normal amount, that it runs parallel to ammonia 
and can be diminished by alkaline treatment ; that the normal ratio between 
the intestinal and the renal ehminations is reversed in favor of the latter 
and that there seems to be a retention of magnesium in the body. In cases 
of arteriosclerosis a retention of calcium has been demonstrated. In acute 
lobar pneumonia Peabody 37 found that calcium was retained but that the 
output of magnesium was normal or so increased that there is a definite 
loss to the body. 

The relation of calcium to phosphaturia is interesting, since this metal 
would appear to be more to blame for this symptom-complex than is the 
phosphoric acid (see page 101). 

Quantitative Determination of Calcium. — Of the filtered urine 
200 c.c. are made alkaline with ammonia until a distinct precipitate is 
visible. This is then dissolved in the smallest possible amount of hydro- 
chloric acid together with the addition of some NaAc. Ammonium oxalate 
is then added in excess and the fluid allowed to stand in the covered beaker 
on a water-bath for 12 hours. Bacterial fermentation should be prevented 
by the addition of thymol or carbolic acid. One thus avoids a precipitate 
of a calcium phosphate which would fall if the ammonia had not been 
added or if the urine becomes foul. After the 12 hours the supernatant 
fluid is decanted through a small ashless filter, the precipitate washed 
Cl-free by decantation with hot water and then finally brought onto the 
paper. The precipitate of CaO is very fine and apt to pass through the 
paper, hence is washed as much as possible by decanting. The washwater 
may be saved for the determination of magnesium. The filter paper is 
then dried and put into a platinum dish, burned slowly for a long time, then 
at a dull red till the mass on cooling is perfectly white. Since this ash will 
still contain some oxide of calcium it is moistened with a concentrated 
solution of ammonium carbonate, dried slowly and very gently ignited. 
This treatment with ammonium carbonate is repeated till constant weight 
as calcium carbonate is reached. One part of CaCO 3 equals 0.40 parts of Ca. 

Another method is to burn the mass white with a blast-flame. The 

37 Jour. Exp. Med., January 1, 1913, vol. xvii, p. 71. 



142 CLINICAL DIAGNOSIS 

crucible is then cooled, weighed and the blast repeated until the weight is 
constant. The precipitate is now CaO, i part equalling 1.845 °f calcium 
phosphate. Or, the precipitate is burned white and the concentrated am- 
monium sulphate then added and it is again burned and this repeated until 
there is no increase in weight. One part of the calcium sulphate ash equals 
0.41 1 76 part of CaO. 

Quantitative Determination of Magnesium. — For determination 
of magnesium the filtrate and washwater of the calcium determination 
may be used. One-third volume of 10% NH 4 OH (sp. gr. 0.96) is added 
which will precipitate all of the Mg as NH 4 MgP0 4 . This precipitate is 
allowed to settle well, is collected on an ashless filter, washed with water 
plus % volume of ammonia, dried thoroughly, shaken into a platinum cru- 
cible, the paper burned in a platinum spiral and its ash added to the cru- 
cible. The whole is then fused. Since the precipitate will contain some 
uric acid it will not burn quite white. It should, therefore, be cooled, a 
small piece of NH4NO3 and a few drops of water added, warmed slowly 
and then finally burned. The result is Mg 2 P20 7 . One hundred parts of 
this equal 36,208 parts of MgO. 

Another good method is to treat 200 c.c. of the original urine in this 
way, which will determine the calcium and magnesium together. The 
calcium alone is then determined in a second portion. The difference will 
be the magnesium. 

Sodium and Potassium. — The daily urine contains from 4.2 to 7.4 gms. 
of sodium weighed as Na 2 and from 2.3 to 3.9 gms. of potassium weighed 
as K 2 0. The usual relation between them is 5 13. 

The amount of these alkalies present will depend in general on the food. 
During hunger periods and also in fever the potassium may exceed the 
sodium but after the crisis the sodium will again predominate. 

Severe exercise and also a vegetable diet will increase the amount 
of potassium. 

Iron. — A trace of iron in organic combination is always present in the 
urine. It would be of great value could we determine this accurately, but 
unfortunately the methods used have too many sources of error. The 
reagents, for example, may contain more iron than the specimen to be 
examined. The amounts of urine-iron claimed as normal for 24 hours 
vary from 1 to 10 mg. It has been found increased in fevers, in malaria 
(even 16 mgms.) in pernicious anemia and in alcoholism. Neumann and 
Mayer, 38 using Neumann's method of ashing the urine, found that the daily 
output of iron of a normal person varied from 0.93 to 1.139 mgms. (aver- 
age 0.983 mgm.). They found it increased especially in the urine of 
alcoholics and made the interesting observation that in diabetes mellitus 
the output of iron runs parallel to that of the sugar, being quite con- 
stantly 2.5 mgms. of iron per 100 gms. of sugar. 

38 Zeitsch. f. physiol. Chem., 1902, vol. xxxvii, p. 2. 



THE URINE: PIGMENTS 143 

Lead. — To test urine for lead a considerable volume is -evaporated to 
dryness and 50 c.c. of fuming HN0 3 added to the residue. After the reac- 
tion has subsided this is allowed to simmer over the free flame for l / 2 hour 
and 25 c.c. more of acid are added. This is repeated 3 times, each for 15 
minutes. The resulting fluid is then evaporated to small volume, is neutral- 
ized with NaOH, filtered and is tested for lead with H 2 S. If lead is present 
a brown precipitate will form. 

Arsenic may be detected by saturating the faintly acid urine with H 2 S, 
allowing it to stand for from 12 to 24 hours, then filtering, washing and 
treating the precipitate with bromine water, which will dissolve any arsenic 
sulphide. This solution is transferred to a suitable flask, zinc and sulphuric 
acid are added and the resulting stream of hydrogen is conducted into an acid 
AgN0 3 solution (AgN0 3 0.1 to 0.2 gm. ; HN0 3 2 gms. ; water 10 c.c). If AsH 3 
is generated a blackish-brown precipitate of metallic arsenic is the result. 

PIGMENTS OF THE URINE 

Indoxyl Sulphate. — The value of the ethereal compounds of sulphuric, 
glycuronic and other acids, usually referred to as " indican," has been 
overestimated, yet they do have a certain clinical importance. Indoxyl 
sulphate, the chief, i.e., the one of the " indican " bodies easiest to demon- 
strate, originates in the intestine as indol, a decomposition product of 
proteid. This is absorbed from the intestine, in the body is oxidized to 
indoxyl, is conjugated, with sulphuric acid and is excreted as an alkaline 
salt. None is present in the urine of a new-born child or even before it is fed 
cow's milk. In adults on mixed diet a certain amount (from 5 to 25 mgms. 
in 24 hours), is always present. In man its output is greater on a flesh 
than on a vegetable diet. That present in the urine of a fasting person 
arises from the decomposition of the intestinal secretions. To be of value 
chemically this body must be present in the urine in considerable amounts. 
Often it does reach from 50 to 150 mgms. in 24 hours. It should at this 
point be emphasized that an increase of indoxyl and one of the ethereal 
sulphates are not synonymous. We usually estimate the amounts of 
both by the color tests of indoxyl but this may be greatly increased 
when the total ethereal sulphates are not and when the latter are 
increased only a relatively small amount may be bound to indoxyl. 

In general the output of indoxyl sulphate (as indoxyl) is increased by 
the rapid decomposition of proteid either in the lumen of the intestine or 
elsewhere in the body. Its increase in cases of peritonitis and ileus is 
important, since its formation seems to depend upon the presence of trypsin. 
In paresis (from peritonitis or obstruction) of the small intestine the output 
of indoxyl shows a great and rapid increase ; in paresis of the colon there is 
either no increase or one which begins late and is due perhaps to the bac- 
terial decomposition of food. The determination of indoxyl would not help 
to differentiate between peritonitis and intestinal obstruction involving 



144 CLINICAL DIAGNOSIS 

the same portion of the bowel. In one very interesting case of syphilitic 
stricture of the ileum the frequent attacks of partial obstruction due to 
the food could be accurately foretold by the increase of this body in the 
urine. It is much increased in cases of intussusception, of new growths 
and of twists of the small intestine. It is increased by intestinal putre- 
faction due to any cause, especially that present in the cholera infantum 
of children, in typhoid fever, in dilated stomach and some cases of nephritis. 
In these conditions a brisk purging will diminish its output greatly. 

It is increased by the decomposition of albumin anywhere in the body ; 
in gangrene of the lung, fetid empyema, putrid bronchitis (in which cases 
it may be present in very large amounts) and in advanced pulmonary or 
intestinal tuberculosis. Coriat thought its increase i element of the symp- 
tom-complex of akinetic mental conditions and its diminution i element 
of that of the hyperkinetic states. He considers this fluctuation in these 
patients not due to any intestinal condition nor to the diet. In certain 
cases of chronic constipation the urine may contain indoxyl in large quan- 
tities, but not necessarily much ethereal sulphates. One such case, a 
colleague of mine who enjoyed the best of health, furnished my classes for 
several years with urine very rich in this pigment. He gives a history of 
some severe abdominal condition when ten years of age, since which time 
he has been troubled with constipation. 

The following is the analysis of his urine : 

Total amount, in 24 hours, 1770 c.c. Color, clear yellow. On boiling, 
the color becomes dark brownish-red, almost black, with a dark magenta 
foam. Beautiful indoxyl test. 

Total SO3, 1.59 gms., of which the ethereal sulphates were only 14%. 
Total sulphur, 1.82 gms. (as SO 3) in 24 hours. 

The urine gave a splendid indigo-blue but not Rosenbach's test. 

It will be seen that despite the color on boiling and the good indoxyl 
test the ethereal sulphates of this specimen were not increased. 

Its output is always diminished by closure of the pancreatic duct, 
but such closure cannot be assumed when there is absence of indoxyl 
sulphate unless other evidence is positive. It is increased when the HC1 
of the gastric juice is diminished. In nephritis it would seem to bear some 
relation to the albumin output but this needs confirmation. 

Indigo calculi have been found. 

The demonstration of indoxyl and the estimation of its amount depend 
on its oxidation to indigo-blue. 

2C 8 H6NKS04+02 = 2C8H 5 NO+2KHS04 

This reaction occurs if an oxidizing agent be added, or when the urine 
decomposes, in which case indigo may collect on the surface as a copper-red 
scum with a metallic glistening. Rarely, however, is enough present to 
be seen grossly. 



THE URINE: PIGMENTS 145 

Indigo-blue is a dark blue powder, insoluble in water, slightly so in 
chloroform but easily soluble in hot aniline. It is insoluble in alcohol 
and ether. It should be collected on an asbestos filter, washed with water 
then with alcohol (to separate the indigo-red) and then dried. 

Indigo-blue may be sublimed at 300 C, giving off a purple-red vapor 
which cools in prismatic crystals of a copper-red, metallic color, but deep 
blue by transmitted light. If indigo-blue be mixed with hot alcohol and 
some very strong NaOH and some glucose be added and this fluid fill a 
closed flask, indigo-white is formed. When this is exposed to the air indigo- 
blue will recrystallize out. 

Tests of Indoxyl Sulphate — Jaffe's Test. — Albumin must be first 
removed from the urine by boiling and filtering. One test-tube is half 
filled with urine and another of the same size with the same amount of 
concentrated HC1. On the lips of this latter test-tube is placed the smallest 
possible drop of fresh concentrated Ca(C10) 2 . The HC1 is then poured 
quickly into the tube containing the urine, carrying with it as it flows over 
the edge this drop of hypochlorite. The fluids are now mixed rapidly by 
inverting (not by shaking) the tube. One cubic centimeter or so of chloro- 
form is then added to extract the indigo as it is formed. If necessary another 
drop of the Ca(C10) 2 solution may be added after the fluids are mixed. 

This test may be performed also as a contact test. 

Hammersten advises to add to 20 c.c. of urine 2 or 3 c.c. of chloroform, 
an equal amount of hydrochloric acid, and then at once the Ca(C10) 2 
solution drop by drop, reversing the tube several times after each addition. 
The difficulty with this test is that a slight excess of the hypochlorite will 
destroy the indigo, giving yellow isatin." 

Ca(C10) 2 is a difficult substance to obtain pure, since it deteriorates 
so rapidly that the manufacturing chemists refuse to handle it. A pure 
salt is unnecessary, and the ordinary cheap bleaching powder or " chloride 
of lime " is satisfactory. (Chloride of lime is not calcium chloride, but a 
mixture of calcium hydroxide, calcium chloride, and calcium hypochlorite.) 

Obermayer's Test. — The urine is first freed of disturbing substances by 
precipitating it with about one-fifth its volume of 20% PbAc, avoiding an 
excess, and then filtering. An equal amount of fuming hydrochloric acid 
containing a little Fe 2 Cl 6 (4 parts of Fe 2 Cl 6 in 1 liter of HC1) is then added. 
In a few minutes the blue color appears. The indigo can be extracted 
with chloroform. 

If the urine contains potassium iodide a violet color results ; if thymol, 
a bluish-green. 

Since indol may disturb a bile test with HN0 3 all urines which contain 
indoxyl should first be extracted with acidulated chloroform before they 
are tested for bile. 

Indigo if evaporated from chloroform will crystallize out in needles 
or plates. 
10 



146 CLINICAL DIAGNOSIS 

Quantitative Determinations. — The quantitative determinations of 
indoxyl are in general unsatisfactory, and workers usually are satisfied with 
approximate estimations from the depth of color obtained by Obermayer's 
reagent. A more accurate result is obtained by repeatedly extracting the 
urine and evaporating the extracts in a weighed beaker. All indigo-red 
should first he removed by washing the residue with alcohol. The residue 
is then dried at from 105 to 11 o° C, and weighed. 

Ellinger 39 found that but 85% of the amount of indigo theoretically 
present could be obtained in this way (evidently isatin is formed) , and that 
neither the concentration nor the excess of reagent are of moment if the 
indigo-blue is extracted quickly enough. 

Another method is to make the urine slightly acid if necessary with 
acetic acid, precipitate it with Xo its volume of PbAc and, if it is concen- 
trated, dilute it Y 2 with water. To a measured portion of the filtrate is 
then added an equal volume of Obermayer's reagent. It is then shaken 
out several times with chloroform until this is no longer colored. (The 
amount of filtrate chosen should be such that 3 to 4 extractions for 2 
minutes each time with 30 c.c. of chloroform are enough.) The filtered 
extract is distilled, the extract dried for 5 minutes then washed out 2 to 3 
times with hot water (to remove isatin), dissolved in 10 c.c. of concentrated 
H2SO4, this solution diluted to 100 c.c. with water and titrated with a 
dilute KMn04 solution (5 c.c. of a 0.3% solution diluted to 200 c.c.) which 
has been standardized against pure indigo-blue. About 87% of the correct 
amount can in this way be measured hence in using this test a correction 
of % should be made. A double determination by this method requires 
about one and a half hours. 

Strauss also used Obermayer's solution and extracted the indigo 
v.ith chloroform, using a small separating funnel similar to that used in 
the lactic acid test (see Fig. 70). The combined chloroform extracts are 
measured, 2 c.c. are then removed and diluted till its color matches that 
of a standard tube of known content and from this can be reckoned the 
total amount. 

Coriat proposed i0 a graduated separating funnel, in which the Ca(ClO) 2 
test is made and the chloroform extract compared with a standard color. 

Phenol is almost always increased in the urine when the output of 
indoxyl is, but the reverse is not true. 

A urine rich in indoxyl is as a rule normal in color when voided. In 
certain cases, however, the oxidation has already occurred and the urine 
is green when voided (green because of the blue of the indigo and the 
yellow of the urine) . Such cases have been described by Sahli, MacPhedran 
and Goldie, and by others, but must be excessively rare and methylene 
blue should always be excluded (see page 99). 

39 Zeitschr. f. physiol. Chem., vol. xxxviii, p. 178. 

40 Am. Jour. Med. Sci., April, 1902. 



THE URINE: PIGMENTS 147 

Skatoxyl-Sulphate. — Skatol, another product of the bacterial decom- 
position of albumin, also is formed in the intestine and absorbed. By 
analogy we may suppose that like indol it is oxidized to skatoxyl, conjugated 
with sulphuric acid and eliminated by the urine. As a matter of fact, 
however skatoxyl sulphuric acid has never been actually demonstrated 
in the urine and the colors which would suggest its presence may as well 
be due to other red pigments. 

The red or violet color produced by adding to the urine an oxidizing 
agent with a strong acid is usually attributed to skatol-red. With Fe 2 Cl 6 it 
gives a violet color and with concentrated HO it is decomposed, depositing 
a red precipitate. In Jaffe's test the urine becomes dark red or violet. On 
standing -exposed to the air such urines darken from above downward, 
first red, then violet, finally even black. But as has already been mentioned, 
th Q se colors are not conclusive for skatol-red. Rosin denies that skatoxyl 
has ever been proven present and thinks that all these tests could be 
explained by indigo-red. To prove that a pigment is skatoxyl it would 
be necessary to reduce it with zinc-dust and obtain skatol. 

Indigo-red. — The pigment called indigo-red has several other names, 
e.g., urorubin and urorhodin. This body is always formed with indigo-blue 
especially if Jaffe's test be made with warm urine. Indigo-blue and indigo- 
red are isomeres which arise from the same mother substance (indoxyl 
sulphate) . Indigo-red forms spontaneously in decomposing urine and may 
form a sediment. Urorosein is formed at the same time. 

Indigo-red crystallizes in dark reddish-brown or chocolate-brown 
needles or plates. Heated to 295 or 310 C. it sublimes with violet-red 
fumes. It is insoluble in water, dilute acids, and in alkalies. It gives a 
cherry-red solution with alcohol, ether, chloroform, and especially with 
glacial acetic acid. From dilute alcoholic solution it precipitates in crystals. 
From glacial acetic acid it is precipitated by soda or by water. It has a 
characteristic absorption spectrum. 

Reduction Test. — The alcohol solution is made alkaline with sodium 
carbonate, a little glucose added and it is gently warmed. The solution 
decolorizes, but the color returns on shaking it in the air. This can be 
repeated as often as desired. If the pigment be boiled with even dilute 
caustic alkali the red is destroyed and various brown decomposition 
products result. 

This pigment is present in large amounts in certain urines which give 
Rosenbach's test, but alone it is not responsible for the Burgundy-red color. 
It is increased especially in intestinal troubles; ileus, obstruction, cancer, 
etc. It is also present in large amounts in some urines which do not give 
a characteristic Rosenbach test, but these conditions are so various that 
they cannot be classified. It is found in traces in normal urine. 

Demonstration. — Nitric acid added to a urine containing indigo-red 
gives a red color. A great deal of indigo-red is formed by Jaffe's test, 



148 CLINICAL DIAGNOSIS 

especially if the urine be heated. After the urine is cold, it may be neutral- 
ized with soda and then shaken out with ether. The ether takes a fine 
red color and gives the absorption spectrum of this body. The ether extract 
may be evaporated in a watch glass and the crystals obtained. 

Indigo-red is present in certain freshly voided urines, as in cases of 
pyelocystitis. It has also been found in concretions. 

Among other red pigments of the urine is urorosein which is found in 
normal urine. This is characterized by its easy solubility in amyl alcohol 
and its insolubility in chloroform, ether and benzol. Ammonia and alkaline 
carbonates decolorize it at once while acid will restore the color. It has a 
characteristic spectrum. It is very unstable and decomposes rapidly. 

To demonstrate urorosein one adds to the urine %o its volume of HC1 
and then filters. Urorosein makes a red stain on the filter paper. Uro- 
rosein also is produced by Jaffe's test, but is not extracted by the chloroform 
or ether. 

The urine may contain other red pigments and after it is boiled with 
acid various brown pigments are produced to which beginners ascribe 
clinical value but which have none. 

Paracresol and Phenolsulphuric Acid. — The daily urine contains 
from 17 to 51 mgms. of phenol and usually more paracresol. The sum of 
these 2 varies in various conditions and in general runs parallel to the out- 
put of indoxyl. The phenol is increased whenever indoxyl is, but the 
reverse is not always true. They are increased by a vegetable diet, in 
ileus and peritonitis, also in diphtheria, scarlet fever and erysipelas. Little 
is present in typhoid fever, smallpox and meningitis. They are products 
of decomposition which may take place anywhere in the body but especially 
in the intestine. 

Pyrocatechin (see page 98) is a similar body also eliminated conju- 
gated with sulphuric acid, as also is 

Hydrochinon (see page 98) especially after carbolic acid poisoning. 

Potassium iodide of course often appears in the urine and must be 
excluded when testing for pigments. If HN0 3 and then chloroform are 
added to a urine containing KI the latter will take the pink color of iodine. 
Or, after the HN0 3 , powdered starch may be added which will take a 
distinct blue color. 

Bile Pigments in the Urine. — Bile pigment never appears in normal 
human urine (although it does in that of some animals). Bilirubin is 
derived from hemoglobin and so may be demonstrated in the plasma, 
and therefore in the urine, when there is increased breaking down of red 
blood-cells — e.g., in the hemoglobinemia due to a blood poison. Bilirubin 
is very similar to hematin and is an isomer of hematoporphyrin. 

The cases of jaundice have been divided into 2 groups; the " hepato- 
genous," due to complete obstruction of the bile passages, in which bile 
appears at once in the urine, as in cases of catarrhal jaundice, calculus in 



THE URINE: PIGMENTS 149 

the common duct, cancer, or cirrhosis of the liver ; and the " hematogenous " 
jaundice, formerly supposed to be the direct result of hemolysis by poisons, 
as the toxins of severe infections. In these cases bile will appear in the 
urine even before the skin or conjunctivas are stained. Hematogenous 
jaundice was formerly explained as due to the inability of the liver to 
warehouse all of the free hemoglobin, but the correct explanation would 
seem to be that the bile capillaries are choked by the greatly increased 
amount of bile pigment which renders the bile too viscid to flow well 
through the bile ducts. " Toxemic jaundice " has been proposed as a 
better term for this condition. 

The bile pigments and their derivatives of interest in clinical chemistry 
are bilirubin, biliverdin, bilifuscin, biliprasin, cholecyanin, and choletelin, 
all of which are products of bilirubin. Bilirubin is the only one which has 
been demonstrated in fresh urine. Biliverdin is often found in stale urine 
after the bacteria have had time to oxidize some of the bilirubin. The 
other derivatives explain some of the colors in Gmelin's test. It should be 
remembered, that all of the bile pigment present in a specimen of urine 
may be contained in the urate sediment. 

Bilirubin, C32H3 6 N 4 6 . — This is the one pigment of fresh human bile. 
It certainly can arise from hemoglobin elsewhere than in the liver. Its 
calcium salts occur in gall-stones; its crystals, the so-called hematodin 
crystals are met with in old blood extravasations. It is found in the fluid 
of certain cysts, especially of the breast and of the thyroid. 

Bilirubin may be separated from mixtures of pigments by precipitating 
the fluid containing them with milk of lime in moderate amount while 
shaking well, saturating with C0 2 at once to prevent the decomposition 
of methemoglobrn et at, and filtering. The precipitate is washed, dissolved 
in alcohol, chloroform is then added and then acetic acid to separate out 
the calcium. This fluid is filtered, the chloroform separated by adding 
water and the chloroform extract filtered through a dry paper and evapor- 
ated. The bilirubin of the residue is washed with a little alcohol and ether. 
The results by this method are always a little too low since calcium does 
not precipitate all the pigment and some is later destroyed. The work 
must be done rapidly. Hematin, if present, would also be isolated, but 
hematin does not occur in the body, except perhaps in the stomach and 
intestinal contents. 

The crystals of bilirubin are rhombs, often with rounded edges, or 
needles which if pure have a beautiful brown-red color. They are perfectly 
insoluble in water, are soluble in alcohol and in chloroform, especially if 
hot, giving these solutions a brownish-red color. It may be precipitated 
unchanged from alcoholic solution by acids, it forms compounds with alka- 
lies which are insoluble in chloroform but are soluble in water. (Hence 
bilirubin may be washed from a chloroform solution by an alkali. In this 
it differs from lutein.) It is precipitated by BaS0 4 and by (NH 4 )2S0 4 . 



150 CLINICAL DIAGNOSIS 

An alkaline solution of bilirubin exposed to the air becomes oxidized to 
green biliverdin. In an alkaline (decomposing) urine, however, this seldom 
occurs since such would soon contain enough (NH^S, to change the 
biliverdin (and bilicyanin) back to bilirubin. Bilirubin itself may disappear 
from an alkaline urine, also from a urine preserved with chloroform. Bili- 
rubin has no absorption spectrum. 

Biliverdin, C 3 2H3 6 N 4 08, occurs in the bile of many animals, but not 
in that of normal men. It is found, however, in the intestine and vomitus, 
and in jaundiced urine which has stood even for a short time. When pure, 
biliverdin is an amorphous, greenish-black powder, insoluble in water, 
ether or chloroform, but easily soluble in alcohol. In its insolubility in 
chloroform and its solubility in alcohol it differs from bilirubin. Its com- 
pounds with the alkalies are soluble to a green or a brownish-green solution. 
It is soluble in concentrated acetic acid and in HC1. The alkaline solution 
has no absorption spectrum, but an alcoholic weakly acid solution shows 
i band. With Gmelin's test it gives the sams color changes as bilirubin. 
It can be reduced to bilirubin. 

Hydrobilirubin, C32H46N2O7, is considered by some to be the same as 
urobilin, but this is denied by the majority of workers. It is produced from 
bilirubin in the lower intestine by reducing bacteria. 

Bilifuscin is a pigment which probably has not yet been isolated pure. 
The substance described (C32H40N4O8) is of an amorphous brown color, 
is soluble in alcohol to a deep brown solution, and in alkali, ammonia and 
dilute NaOH. It is insoluble in water and ether, and nearly soluble in 
chloroform. It is soluble in ether and chloroform if fatty acids are present. 
In the pure state it does not give Gmelin's reaction. Its spectrum is similar 
to that of biliprasin. 

Biliprasin is said by some to be a mixture of bilirubin and bilifuscin, 
by others to be an intermediate stage between bilirubin and biliverdin, 
while still others consider it identical with biliverdin. The formula given 
is C32H44N4O12. Its alcoholic solution has no absorption spectrum, but 
the alkaline solution has. The brown color of its alcoholic alkaline solution 
is the chief point of difference between this and biliverdin. It differs from 
bilifuscin, since if acid be added to the above solution its color changes to 
green. The Gmelin test is of no value in recognizing this pigment. 

Cholecyanin is a product of the oxidization of bile pigments with 
HNO3, PbO, or KMn0 4 . It may be further oxidized to choletelin. It 
gives a characteristic spectrum. Cholecyanin is insoluble in H 2 is soluble 
in alkalies and strong acids. It may be reduced to bilirubin. Its neutral 
or faintly acid solution has a bluish-green or steel-blue color with a beautiful 
red fluorescence. Its alkaline solution has a green color. The only way 
of recognizing it is by its beautiful spectrum. 

Choletelin. — Choletelin is a product of the oxidation of bilirubin 
with HNO3. It is soluble in alcohol, giving a ruby-red solution. Its 



THE URINE: PIGMENTS 151 

dilute solution has a yellowish-red color which does not change with change 
of reaction, as does that of urobilin. Chemically it is similar to urobilin, 
but its absorption spectrum differs, and with ZnCl 2 it gives no fluorescence. 
It is not precipitated by PbAc. It may be identified accurately only by 
the spectrum of its solution made acid with acetic acid. Its spectrum 
should not be confused with that of urobilin. 

The reducible body of Stokvis is a by-product of the complete 
oxidation of bile pigment. It is a substance soluble in water, alcohol, 
alkali, and dilute acid, but not in ether or chloroform. It is not precipi- 
tated by PbAc, but is by PbAc and NH 4 OH. Its characteristic reaction 
is, that if its alkaline solution be boiled with a reducing substance (e.g., 
(NH 4 )2S) the solution becomes a beautiful rose-red color and gives an 
absorption spectrum. If this solution is shaken with air the rose red will 
disappear, but is soon restored with the disappearance of the spectrum. 

The color of a bile-stained urine does not depend alone on the 
amount of bilirubin present, for a pale urine may contain much and a dark 
one, little. In general, the color of a jaundiced urine varies from a dark 
yellow to a brown or even to a greenish-black. An easy and accurate proof 
of the presence of bile is the yellow color of the foam when the urine is 
shaken since that of a very dark non- jaundiced urine will be pure white 
unless much urobilin be present. If a sediment fall in a bile-stained urine it 
will be yellow in color and may contain all the bilirubin of the specimen. 
Bilirubin will stain filter paper yellow. . A jaundiced urine usually contains 
also an excess of urobilin and indoxyl, hence were the bilirubin to be re- 
moved its color would still remain dark. Such urine always contains the 
nucleo-albumin of bile. Heller's test for traces of albumin should not be 
applied to such urines since the oxidized pigment will confuse one. 

Test. — If the urine contains much bile it may be shaken out with chlo- 
roform, the chloroform extract poured off and evaporated, the residue 
taken up again with chloroform and evaporated in a watch glass. Rhombic 
prisms of bilirubin separate out, which are soluble in alkali, which give 
Gmelin's tests and which on exposure to the air become green. 

Gmelin's Test. — The urine in a pipet is superimposed over crude HN0 3 
(sp. gr. at least 1.4) in a test-tube. (The HN0 3 used should be only faintly 
yellow with HN0 2 . The amount of HN0 2 in the HN0 3 may be increased 
by adding a few pine shavings, or diminished by adding a little urea.) 

If bilirubin be present strata of colors will be seen in the urine which 
from above downward are green, blue- violet, red, and just above the HNO3, 
yellow. The green is the essential part of the test, not the other colors. 

If too much HN0 2 is present the whole urine will soon be yellow. 
This test cannot be applied to very dark urines or to urines rich in indican, 
since the blue of the indigo and the yellow of the urine may give a deceptive 
green color, also the ring which forms at the point of puncture of the two 
fluids has a black tone and contains a fine precipitate. If in doubt the 



152 CLINICAL DIAGNOSIS 

urine may be extracted with chloroform and both pigments tested for. 
A violet-red ring may be due to skatoxyl (?). The violet-red color also 
must, according to some, be present else the test might be confused with 
that of lutein, which gives a blue or a bluish-green ring, but in the case of 
urine this pigment will not confuse one. The violet-red color is due to 
skatol (?) and indoxyl. Biliverdin also gives this test. The test may fail 
if the HNO3 contains too much HN0 2 since the green will not be seen. 

Alcoholic solutions cannot be tested, since alcohol alone will give this 
test. If the urines to be tested are to be shaken out with ether, the ether 
must be alcohol-free, and this is not always the case. 

If the urine contains much urobilin the bile test may be unsatisfactory, 
but this seldom happens. To avoid it the urine should be diluted to a 
specific gravity of 1.005, an d then if the green color appears bilirubin 
certainly is present. Some always dilute the urine this much before making 
Gmelin's test since then only the green will appear. 

The lutein of the serum does not disturb one much but methemoglobin 
may. If abundant albumin is present this should be precipitated and this 
precipitate extracted with chloroform since it will carry much bilirubin 
down with it. If but a trace of albumin and much bile are present the 
albumin may be disregarded, indeed it may even improve it, but if only a 
trace of bile and a trace of albumin are present the latter must be precipi- 
tated, dried and extracted. 

The Gmelin test is said to indicate as little as 1 part of bilirubin in 
80,000 of urine. 

After the ingestion of antipyrin the test is said to be positive. 

Rosenbach's test is the best modification of Gmelin's test. Indeed, it 
is far more sensitive than Gmelin's. As much as possible of the urine 
acidified with HC1 is filtered several times through a filter paper on which 
will collect the bile-stained elements of the sediment and therefore the 
most of its bilirubin. The filter is then partially dried with dry filter paper 
and then 1 drop of yellow HN0 3 dropped upon it. Rings will be seen 
about the drop which will present the above-mentioned play of colors, 
the external one being green. If the paper has been allowed to dry, it 
should again be moistened with water dropping on the acid. Instead of 
filter paper Dragendorfl uses a porous porcelain plate. 

The following tests are excellent as a class exercise since they teach 
well the chemistry of bilirubin. A test-tube is filled full of urine, 2 c.c. of 
chloroform and 3 drops of HC1 added and the whole thoroughly mixed. 
The bile pigment, which acts as acid, is set free from its alkali combination 
by the HC1 and being more soluble in this condition in chloroform than in 
water, is extracted by the chloroform. The chloroform is then poured off 
into another test-tube and an equal amount of water added. One drop of 
NaOH will now transform the free pigment to an alkali salt, which is 
soluble in water. The aqueous solution may now be tested with nitric 



THE URINE: PIGMENTS 153 

acid, or the chloroform extract may be evaporated in a watch glass and 
the crystals studied. 

According to another method the urine is rendered alkaline with NaOH 
or soda and then precipitated with BaCl 2 or CaCl 2 , or the hydroxides of 
these metals, as long as a colored precipitate falls. This yellow precipitate 
is then filtered off and boiled with alcohol plus a few drops of dilute H 2 S0 4 . 
If no pigment at all is present this fluid remains colorless; but if bilirubin 
is present a beautiful, clear, green solution is obtained. If the urine con- 
tained chrysophanic acid the fluid becomes orange-yellow. This test is 
positive when others are negative. 

Bilirubin may also be extracted from acid urine with chloroform (add 
a few drops of HC1). To avoid an emulsion, however, do not shake too 
vigorously. In case it be necessary to test the urate sediment, and this 
may contain all of the bile pigment which was in the urine, the sediment 
is dissolved with soda and the solution tested for bilirubin. 

Hammarstens Test. — Hammarsten's reagent consists of i part of 25% 
HNO3 and 19 parts of 25% HC1. This acid mixture is allowed to stand 
until it is yellow. Just before each test 4 parts of alcohol are added to 
1 part of this reagent. If to a few cubic centimeters of this fluid just mixed 
are added a few drops of a pure bilirubin solution a beautiful permanent 
green color is at once obtained. By adding more acid we can get at will 
the other colors, even the yellow. This test is applied to the urine as 
follows : Ten cubic centimeters of the urine to be tested are poured into a 
15 c.c. centrifuge tube, a few cubic centimeters of BaCl 2 or CaCl 2 solution 
added and the mixture centrifugalized for from l / 2 to 1 minute. The super- 
natant fluid is then poured off. To the sediment are then added 1.2 c.c. 
of the above acid reagent, the tube well shaken and centrifugalized for 
half a minute. If CaCl 2 were used, this last centrifugalization is not 
necessary. A green solution is obtained even if there is but the 1 part 
of bilirubin in 500,000 to 1,000,000 parts of urine. 

Hup per? s Test. — Ten cubic centimeters of urine are made alkaline 
with soda and CaCl 2 solution is added as long as a precipitate is formed. 
This is filtered through a small filter and the precipitate washed with 
water. The filter paper with the precipitate is then placed in a porcelain 
dish, acid alcohol (5 c.c. HC1 in 100 c.c. of alcohol) added, and then heated. 
In the presence of bilirubin the color of this fluid becomes green or blue. 
This test is recommended in case the urine contains much indican or is 
quite dark in color. 

Nakayama's Modification of Httpperfs Test. — Nakayama's reagent con- 
sists of 95% alcohol 99 parts, fuming HC1 1 part and Fe 2 Cl 6 4 gms. per 
liter of the above mixture. To 5 c.c. of the urine, acidified if necessary, 
is added an equal amount of 10% BaCl 2 solution and the fluid centrifugal- 
ized. The supernatant clear fluid is then poured off and 2 c.c. of the above 
reagent are added to the precipitate. The fluid is then heated to a boil. 



154 CLINICAL DIAGNOSIS 

A green or a bluish-green solution is obtained which on the addition of 
yellow HN0 3 becomes violet or red. The test is said to indicate 1 part 
of bilirubin in 1,200,000 parts of urine. 

The following very important test together with its modifications has 
been reported under at least 4 different names, Trousseau's, perhaps, having 
priority. The urine, acidified if necessary with acetic acid, is mixed with 
a tincture of iodine ; or a contact test made, the iodine tincture being super- 
imposed upon the urine. CI or Br also may be used instead of I. In the 
presence of bilirubin a fine emerald-green color is obtained which is not 
biliverdin but a substitution product of bilirubin with iodine. This test 
is more sensitive than Gmelin's; it may be made even more delicate if 
the tincture of iodine be diluted 1 : 10 with alcohol (hence a 1% iodine 
solution) and the urine be overlaid with this. A green ring which appears 
at once, or in 1 minute, will indicate bile (Rosin). Using this test there 
can be no confusion with indoxyl. It is said, however, that some normal 
urines will give a positive test. 

Stokvis's Cholecyanin Test. — This test serves as a good control if bile be 
present together with other pigments in excess but it is not as delicate as 
are some of the above. To from 20 to 30 c.c. of urine are added 5 to 10 c.c. 
of ZnAc or of 20% ZnCl 2 solution. A little soda is added to reduce the 
acidity, after which it is filtered. The precipitate which contains all of 
the bile is now dissolved in NH 4 OH. This solution of bile pigment now 
in the form of cholecyanin is next neutralized. It will have a blue-green 
color with a red fluorescence and will give a characteristic 3 -band spectrum. 

Some of these tests are good when bile alone is present, others in the 
presence of other pigments also. 

Certain substances occasionally present in the urine after the use of 
rhubarb, senna and santonin should not be mistaken for bile since these 
urines become red on the addition of an alkali, but the original color is 
restored if the urine be again acidified. 

It is important to recognize the crystals of bilirubin. They are often 
found in a jaundiced urine which has been concentrated for leucin and 
ty rosin. To demonstrate them the urine is rendered acid with HC1 and 
allowed to stand in the cold. Bilirubin will precipitate out in intensely 
brown sheaths or rhombs often with rounded edges; their color should 
prevent any confusion. 

It is sometimes desirable to remove bile from the urine. This may be 
done by extracting with chloroform the urine after it has been acidified 
with HC1 or by boiling it briefly with a little animal charcoal. This latter 
method should be carefully used, since other substances, perhaps the one 
sought for, may also be removed. 

KMn0 4 in acid solution destroys the bile pigments perfectly. For 
each 1 c.c. of urine are added 2 drops of HN0 3 (or of HC1) and 2 drops of 
4% KMn0 4 . The urine is then warmed and gently shaken. 



THE URINE: PIGMENTS 155 

Bouma 41 recommends the following quantitative determination of bile: 
To 10 c.c. of fresh urine are added 2 c.c. of 20% CaCl 2 solution. The 
urine is then almost neutralized with NH 4 OH. The urine, still slightly acid, 
is then centrifugalized, the fluid decanted, the sediment shaken up with 
water and again centrifugalized to wash the sediment. The fluid is now 
entirely decanted and 5 c.c. of a mixture of 4 c.c. of absolute alcohol and 
1 c.c. of Obermayer's reagent (1.5 gms. of Fe 2 Cl 6 in 1 liter of HC1, sp. gr. 
1. 1 5) are added to the sediment. This is then poured into a test-tube and 
compared with a set of 6 standard tubes to match the biliverdin which 
has been formed. If much bilirubin (more than 100 mgms.) be present the 
urine is diluted with normal urine (thus not diluting the phosphates) and 
the determination repeated. 

Melanin-Melanogen. — In the urine of patients with melanotic tumors 
melanin or melanogen (Morner) may be observed. The chromogen is 
colorless but the urine on standing, or after the addition of an alkali or 
oxidizing agent, turns black, beginning at the top. This change of color 
may be intensified by adding HN0 3 or Fe 2 Cl 6 . This pigment is insoluble 
in chloroform, which prevents its confusion with indoxyl. It may settle 
out as an amorphous sediment. It is decolorized by boiling with HN0 3 . 

Rosenbach's Reaction. — If to certain urines, while boiling, strong nitric 
acid is added slowly drop by drop, their color will change to a Burgundy 
red and the foam takes on a bluish-red color. The color of the foam is the 
more important part of the test since the red of the urine may be due to 
urobilin. (If an excess of HN0 3 was used the color of the urine would be 
a yellowish-red or yellow and the foam yellow.) If soda or ammonia is 
next added drop by drop the result will be a bluish-red precipitate soluble 
in excess to a brownish-red solution. This test is said to be due to indigo- 
red (Rosin). It has the same significance as the indoxyl reaction. 

Bile Acids. — Glycocholic and taurocholic acids are, contrary to a 
former belief, not present in normal urine, but they may be present in large 
amounts in the urine of patients with jaundice, especially of the obstruc- 
tive type, although, even in these cases, the urine may contain almost 
none and in cases of toxic jaundice they appear but in traces. Their pres- 
ence, however, has but little value in the differential diagnosis between 
obstructive and toxic jaundice. Their detection in the urine is impossible 
unless at least 0.5% is present. 

To isolate the bile acids Thierfelder recommends that the urine be con- 
centrated to a small volume, the residue extracted with strong alcohol 
and then filtered. The alcohol is then removed by evaporation and the 
residual fluid precipitated with basic PbAc and NH 4 OH. This precipitate 
is brought upon a filter paper, is washed with water, dried, treated with 
boiling alcohol several time« and filtered hot. To this filtrate are then 
added a few drops of soda solution to decompose the Pb salts. It is then 

41 Deutsch. me l. Wochenschr., 1904, No. 24. 



156 CLINICAL DIAGNOSIS 

evaporated to dryness, the residue extracted with absolute alcohol and 
filtered. To the filtrate is then added a great excess of ether. It is now 
allowed to stand in order that the sodium salts of the bile acids may pre- 
cipitate out. These at first will be amorphous but later crystalline. The 
precipitate is dissolved in water and tested by Pettinkofer's test. 

Tyson Method. — From 180 to 240 c.c. of urine are evaporated to dryness 
on the water-bath, an excess of absolute alcohol added to the residue and 
this then filtered. To this filtrate are added from 12 to 14 volumes of ether 
which will precipitate the bile acids. This precipitate is then brought 
upon the filter paper, dissolved in distilled water and decolorized with 
animal charcoal 

Pettinkofer's Reaction. — To a solution of bile acids in a test-tube are 
added a little cane-sugar and then slowly, drop by drop, concentrated 
H2SO4, shaking the fluid well all the time and keeping its temperature at 
about 70 C. by warming or cooling as the case may be. A precipitate of 
cholic acid forms first which soon redissolves. Then, as more H 2 S0 4 is 
added, the color of the fluid becomes first cherry red and later a beautiful 
purple which in 8 days will change to a bluish-red. This play of colors is 
due to the reaction of cholic acid and the furfurol which is formed by the 
action of sulphuric acid on cane-sugar. The purple-red solution should be 
diluted with alcohol and examined spectroscopically. It has a character- 
istic absorption spectrum. This is necessary in order to exclude certain 
confusing substances which may be present, as albuminous bodies, many 
bodies easily decomposed by H 2 S0 4 , many pigments, amyl alcohol and 
oleic acid. 

Udrdnzky's Test. — According to Udranzky's test, which is the best, 
1 drop of a 0.1% watery furfurol solution is added to 1 c.c. of the solution 
to be tested. This is underlaid with 1 c.c. of concentrated H 2 S0 4 , the tube 
meanwhile being cooled to restrain the reaction. In the presence of but 
0.033 mgm. of cholic acid one may get a positive test, i.e., a red color, with a 
definite bluish tinge, which on standing becomes blood-red. If 0.05 mgm. 
is present the solution will give a distinct absorption line in the spectrum. 
The spectrum must always be examined for confirmation. A 10% cane- 
sugar solution will give as good a test as a 0.1% furfurol solution. 

If cane-sugar is used in excess it will be burned, giving a brown or a 
black color. An excess of furfurol would give an orange color. Oxidizing 
bodies if present will prevent the reaction. Indoxyl gives a violet color. 
Since some concentrated normal urines give beautiful positive reactions, 
to avoid mistake the bile acids must be isolated as lead salts and then 
tested pure. 

Hays test for bile acids is certainly very easy and is said to be even 
more sensitive and accurate than Pettinkofer's test. On the surface of 
the urine (which has been cooled, if necessary, to a temperature not higher 
than 1 7 C.) is sprinkled a little finely powdered sulphur. If the sulphur 



THE URINE: PIGMENTS 157 

sinks at once it indicates a bile acid content of i : 10,000. If it sinks after 
shaking gently and waiting for 1 minute, 1 : 40,000. It is positive if even 
but 1 : 120,000 of bile acid is present. The explanation given for this 
phenomenon is that bile salts lower the surface tension of the urine. 42 

Diazo Test. — Certain diazo bodies combined with aromatic compounds 
give a colored reaction. A urine test depending on this reaction is recom- 
mended by Ehrlich for clinical use. What body or bodies in the urine give 
it are unknown, but the empirical value of the test is beyond dispute if 
the technic of Ehrlich is accurately followed : 

Fluids: (1) One-half per cent. NaN0 2 . This should be quite fresh. 

(2) Five parts of sulphanilic acid, 50 of HC1 and 1000 of distilled water. 

To 250 c.c. of the second reagent are added 5 c.c. of the first. Only a 
fresh mixture (not over 1 day old) should be used. Equal parts of the 
urine and this mixed reagent are shaken together until considerable foam 
is produced and ammonia is then quickly added in excess. Usually it is 
added drop by drop notwithstanding the warnings not to modify the orig- 
inal technic. If the test be positive the urine will take an intense red and 
the foam a more or less brilliant rose-red color. A brown color or a salmon 
tint are not positive. If the test is positive and the tube be allowed to 
stand a precipitate should form, the upper surface of which has a dark 
greenish-black, or violet, color. Some would have us wait 24 hours for 
this precipitate but most consider that the important part of the test is 
the rose-red color of the foam. 

Since with strong enough reagents normal urines give a positive test 
Green recommends that 1 part of solution 1 and 100 of solution 2 be 
used. This renders the test more delicate and gives fewer unexpected 
positive results. 

Zunz prefers the paramido-acetophenol of Friedenwald's formula 43 
to the sulphanilic acid since the results are more delicate and intense 
(paramido-acetophenol, 50 gms., cone. HC1, 50 c.c, water, q.s. ad 1000 c.c). 

Four drops of a 0.5% solution of NaN0 2 and 10 c.c. of the above solu- 
tion are mixed and shaken well with 10 c.c of urine. About 3 c.c of am- 
monia are now added and the color of the foam observed. Zunz adds the 
ammonia all at once, not drop by drop. He considers the color of the 
foam as the important, and the precipitate as the less important, part 
of the reaction. Disturbing bodies can for the most part be removed by 
shaking the urine out with amyl alcohol, and then removing by heating 
it on the water-bath. 

The urine soon loses its property of giving a positive diazo test, but 
after a few days of ammoniacal fermentation the test may reappear. 

If necessary to keep the urine several days before testing it, ether may 
be added. 

42 Beddard and Pembrey, Brit. Med. Jour., Match 22, 1902. 

43 New York Med. Jour., 1894, p. 745. 



158 CLINICAL DIAGNOSIS 

Some claim that to concentrate the urine on a water-bath to a syrup 
(Michaelis) will give a positive test although the fresh urine was negative. 
Zunz did the most of his careful work with such concentrated urines. 
That this does not always help matters was shown by Imhoff who found 
that some concentrated urine of rabbits with experimental tuberculosis 
gives a brown foam, but if diluted to its previous volume the foam is a 
brilliant red. Similar observations were made by Dr. Hirschfelder, who 
tested the undiluted and the diluted urine of patients as a routine. We 
have been in the habit of testing the fresh, the diluted and the concen- 
trated urines. I am told that certain urines giving a negative test 
according to the usual technic will give a positive one if only }{ volume of 
reagent is used. Since a positive reaction may be due to impurities in 
the reagents (e.g., in the ammonia) one should always control the test, 
using distilled water instead of urine. 

What it is which gives the red color when combined with a diazo body 
is not known. Bondzinski found alloxyproteinic acid in all normal urines 
and since this will give a positive test he suggests it as the cause. Clemens, 
however, claims that the body giving the diazo test is sulphur-free. 

Occurrence. — The urine of normal persons never gives a positive test 
if dilute reagents are used. Ehrlich classified the diseases in which it may 
be positive into 4 groups: a. N on- febrile diseases, such as advanced heart 
disease, chronic hepatitis, carcinoma especially of the pylorus, leukemia, 
marasmus senilis, malarial cachexia, tuberculous abscess, etc. In all of 
these it is rarely positive. 

b. Febrile Diseases. — (1) Those in which the test is almost never posi- 
tive, e.g., acute articular rheumatism and meningitis. 

(2) Those in which it may or may not be positive, as pneumonia, scarlet 
fever, diphtheria, erysipelas and phthisis. 

(3) Those in which it is very often positive, as typhoid fever and 
measles. 

Lobligeois found the test positive in 42 of 52 cases of scarlet fever and 
in but 3 of 137 cases of diphtheria. It may therefore have some value in 
the diagnoses of cases of diphtheria with a scarlatinal rash. Brunschwig 
found that in children the reaction is always positive in typhoid, often in 
scarlet fever, quite often in measles, rarely in pneumonia and never 
in whooping-cough. 

Ehrlich considered that in the first 2 groups of fevers a positive reaction 
means a bad prognosis; that its continued absence speaks strongly against 
the diagnosis of typhoid; and that its reappearance in a case of typhoid 
fever together with recrudescence of the fever speaks in favor of a relapse 
or recrudescence of the typhoid infection rather than to a non-typhoid 
complication or sequela. Long continued use has confirmed the accuracy 
of this test although it is not as important as formerly owing to the advances 
in bacteriology and serology. Its value in typhoid fever is also much 



THE URINE: FERMENTS 159 

lessened by the fact that it is usually negative in the early stage of the 
disease when it would be most helpful. 

In phthisis a positive diazo test suggests a bad prognosis although the 
previous opinion of Michaelis that such cases were always fatal is not 
borne out by the experience of others. Boissiere found it positive in 18 of 
130 severe cases. There is some reason to think that in tuberculosis this 
reaction is due not to the tuberculosis but to a secondary infection. 

It cannot be used to distinguish between typhoid fever and miliary 
tuberculosis since it is so often positive in both conditions. It is positive 
also in puerperal fever and in actinomycosis of the lung. 

The work of Zunz 44 is of particular importance since it seems to have 
been done with exceptional care. His conclusions are that the test is val- 
uable in the early diagnosis of typhoid fever and in the prognosis of tuber- 
culous pneumonia, although in the latter disease a positive reaction does 
not necessarily mean a hopeless prognosis; that it helps in the early diag- 
nosis of measles; that it is in favor of tuberculosis in cases of peritonitis, 
pleurisy, and nephritis ; that it is often present in erysipelas; that if present, 
the prognosis in a case of cancer or sarcoma is immediately grave; that 
in cases of pneumonia and pyothorax (non-tuberculous) the test means 
merely a disturbed metabolism; that in certain cardiac affections it 
speaks in favor of a reserved prognosis; and, in conclusion, that it is a 
useful test, although its value has been much exaggerated. 

Many consider that the ingestion of certain drugs inhibits the test, 
as, for instance, phenol, salol, benzonaphthol, etc. 

Plezl 45 found it positive in typhoid from the middle of the first to the 
end of the third week and in measles before and during the period of erup- 
tion. His suggestion is that apart from these conditions it is positive in 
streptococcus septicemia, which also explains its presence in the angina 
of scarlet fever, in advanced phthisis and other forms of severe tuberculosis. 

We have found the test very valuable, expecially in the diagnosis of 
typhoid fever. In our typhoid cases, however, the test very soon becomes 
negative, evidently the result of the diuresis we encourage. 

For a discussion of Ehrlich's dimethylamidobenzaldehyde reaction the 
reader is referred to Simon's article 46 and for Rosso' s test for typhoid fever 
to the discussion by Neuman and Behrend. 47 

FERMENTS 

Several ferments have been demonstrated in the urine in health and in 
disease and in amounts which depend on the general condition of the 
patient. The most important of these is pepsin. To demonstrate this a 
mass of pure fibrin is covered for several hours with the fresh urine. This 

44 Bull, de l'Acad. roy. de med. de Belgique, ser. iv., t. xiv., p. 553. 

45 Wien. klin. Wochenschr., 1903, No. 31. 

46 Am. Jour. Med. Sci., 1903. 

47 Arch, of Int. Med., April 15, 1913, vol. xi, p. 456. 



160 CLINICAL DIAGNOSIS 

will absorb a great deal of the pepsin. The fibrin is then removed from the 
urine and dropped into a flask containing dilute HCJ, and placed in a 
thermostat. If the fibrin becomes digested pepsin was present. Trypsin, 
it is said, has been found in the urine, but this has not yet been confirmed. 
A diastatic ferment has been demonstrated in some urines and the same is 
true of rennm. It is claimed that the decomposition of urea with the forma- 
tion of ammonia bodies is due to a ferment rather than to bacteria but this 
has never been proven. 

If lipase 48 is present at ah in normal urine it is only in traces. It is 
found in jaundice, perhaps in diabetes mellitus, but especially in those 
conditions accompanied by fat necrosis (in dogs after mechanical injury 
of the pancreas and after tying the pancreatic duct), and in tuberculosis 
during the fibrile periods. 49 

Method (Kastle-Loewenhart) . — Into each of 3 flasks are measured 
5 c.c. of urine. The second flask is then boiled. To the urine in the third 
are added 3 drops of phenolphthalein (1%) and then 0.1N NaOH till 
faintly pink. This amount of alkali, determined by this titration, is then 
added to flasks 1 and 2. To each of the flasks are then added 0.25 c.c. of 
ethylbutyrate and 0.1 c.c. toluene and they are left in a thermostat at 
39 C. for 20 hours. At the end of this period an amount of 0.1N HC1 
which is 0.5 c.c. more than the amount of 0.1N NaOH previously added, 
is measured into each, each flask is then shaken out with 50 c.c. of ether 
and 25 c.c. of alcohol, 3 drops of the phenolphthalein solution are added 
to the ether extract and the amount of butyric acid split off titrated with 
0.1N KOH. 

In case 5 c.c. of urine are not available for each flask the figure obtained 
from the smaller amount is calculated for 5 c.c. using the formula that the 
amount of ferment action varies as the square root of the amount of ferment 
present. 

Amylase of the Urine. — Wohlgemuth's method for the quantitative 
estimation of amylase in the body fluids is as follows : 50 

Ten carefully cleaned and dried test-tubes numbered 1, 2, 3, 4, 5, 6, 7, 
8, 9, 10 are placed in a test-tube stand and to each tube except Tube 1, 
which contains 1 c.c. of the fluid to be tested, is added 1 c.c. of a normal 
saline solution. With a 1 c.c. pipet, accurately graduated to 0.1 c.c, 1 c.c. 
of fluid to be tested is added to Tube 2 , thoroughly mixed drawing up and 
expelling 3 times the mixture of normal saline solution and test fluid. 
Then finally draw up 1 c.c. of the fluid now diluted 1 to 2 and place this 
in Tube 3, repeating the same process of mixing and again withdrawing 
1 c.c. of mixture. This is to be repeated in all the succeeding tubes and 

48 See Hewlett, Jour. Med. Research, 1904, vol. vi., p. 377; also Gamier, Compt. 
rend., 1903, vol. v, p. 1064. 

49 White and Zeedick, M. of the Assoc, of Am. Phys., 1916. 

50 Geyelin, Arch, of Int. Med., 19 14, xiii, p. 96. 



THE URINE: FERMENTS 161 

in the last tube the i ex. withdrawn can be discarded. This will give the 
following amounts of the fluid to be tested in each tube (7 tubes suffice in 
most cases). 

No. 1 = 1.0 c.c; No. 2=0.5 c - c -i No. 3=0.25 c.c; No. 4 = 0.125 c.c; 
No. 5 = 0.0625 c.c; No. 6 = 0.032 c.c; No. 7 =0.016 c.c; No. 8 = 0.008 c.c; 
No. 9 = 0.004 c.c; No. 10 = 0.002 c.c 

Now to each tube are added 2 c.c of 0.1% starch solution (Kahlbaum's 
soluble starch). The tubes are shaken gently and fitted into a wire cage 
which is placed on a water-bath at 38 C. for %. hour. The temperature 
should be kept within >2 degree of 38 C. and the time accurately measured. 
At the end of this time all tubes are withdrawn and placed in ice-water 
for 5 minutes to inhibit further amylolytic activity and are then replaced 
in test-tube stand. To each tube is then added 2 drops of 0.02 $N iodine 
solution which must be freshly prepared daily from a stock solution of 
0.1N strength. One-to-one-thousand starch solution should be freshly 
prepared every 3 days and kept in the refrigerator. 

If the urine is normal the first 2 or 3 tubes will be golden yellow in co'or, 
all starch having been digested, while the fourth will be reddish with no 
tinge of blue, the fifth violet and the others blue. The tube from which 
the reading is taken is the deep-red tube. This color shows that all starch 
has been digested at least to the dextrin stage. 

The calculation is then made as follows: In Tube 4, e.g., there was 
0.125 c - c - urine or fluid which digested 2 c.c. of a 0.1% starch solution in 
}{ hour at 38 G. Therefore 1 c.c. of fluid to be tested would digest starch 
according to the following equation: 

0.125 : 2 c - c - (starch) : : 1 c.c. : X = i6. 

One cubic centimeter of fluid digested 16 c.c. of a 0.1% starch solution 
in Y 2 hour ; this Wohlgemuth designates as follows : 

When 24-hour digestion is employed he expresses it 

D^-=X. 

24 hr. 

The conclusions thus far obtained with this test are interesting although 
not yet valuable. Under normal conditions the d (units of amylase per 
1 c.c. of urine) is almost constant, the lower normal limit being 8, but in 
77.1% of the cases of nephritis is sub-normal or zero. In the other cases 
the urine contained considerable albumin which itself increases enzymatic 
activity or is accompanied in its excretion by the excretion of more amy- 
lase. The d values run fairly parallel with the elimination of phenolsulphone- 
phthalein but in cases of cardiac decomposition the latter usually is sub- 
normal but d normal. 
11 



162 CLINICAL DIAGNOSIS 



CARBOHYDRATES AND ALLIED BODIES IN THE URINE 

Certain carbohydrates in small amount are normal ingredients of the 
urine. Three such have been demonstrated; glucose, animal gum and iso- 
maltose. Bodies related to carbohydrates also may be present in normal 
urine; e.g., the paired glycuronic acid compounds, chondroidin-sulphuric 
acid, nucleinic acid, the mucoid, of: the nubecula and, sometimes, pentose. 
The total output of all of these carbohydrates measured as glucose amounts 
to from 2 to 2.23 gms. in 24 hours. Of glucose itself the normal urine may 
contain from 0.38 to 0.62 gm. in 24 hours (Naunyn, 0.4 to 1.4 gms.). The 
latest report is that of Myers 51 who concluded that the urine of normal 
persons may contain from 0.08 to 0.2% of sugar. 

The total carbohydrates of the urine, both the fermentable and the 
unfermen table, may be determined as benzoylesters. The urine is made 
alkaline with NaOH and the phosphates removed by nitration. To the 
nitrate in a flask is added benzoylchloride, 4 c.c. per 100 c.c. of urine, and 
40 c.c. of 10% NaOH. The flask is now shaken gently for 10 minutes 
(to avoid emulsion), then vigorously for 20 to 25 minutes, until all odor 
of the benzoylchloride has disappeared. It is then allowed to stand for a 
few hours, but not over night since the precipitate would get sticky and 
not filter well, then filtered, the precipitate washed, dried over H2SO4 
and weighed. 

While a normal person should be able to digest, warehouse and use an 
indefinite amount of starch ingested in 1 meal without the appearance 
of sugar in the urine, yet in the case of sugar there is for every person a 
limit to the amount which can be thus ingested without producing a gly- 
curesis. The largest amount of glucose which a person can ingest without 
glycuresis is called his assimilation limit. For the normal person this is 
about 200 gms. (some say 300 gms.). Any amount beyond this produces a 
slight glycosuria of seldom more than 1 gram of glucose. Such a glycosuria 
is termed an alimentary glycosuria, or an alimentary glycosuria e saccharo. 
This follows directly the ingestion of the sugar and lasts but a few hours. 
A glycosuria which follows a meal rich in starch is called an alimentary 
glycosuria e amylo. This indicates a worse injury to the carbohydrate 
metabolism than does a glycosuria of the e saccharo group, and is practically 
always diabetic. There are on the other hand conditions (e.g., pituitary 
disease) which raise the assimilation limit so that a person may consume 
even 500 or more grams of glucose and remain sugar-free. For each sugar 
there is a different assimilation limit. Hofmeister found that galactose 
and lactose passed the most readily into the urine, while for dextrose, 
levulose and cane sugar the limit is much higher. 

A spontaneous glycosuria is one which appears while the patient is on 
an ordinary mixed diet ; that is, a diet which does not contain an unusual 

51 Proc. Soc. Exp. Biol, and Med., 1916, xiii, p. 178. U.) 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 163 

amount of carbohydrate. A temporary spontaneous glycosuria is one 
which lasts but a few days. A spontaneous glycosuria which continues 
for several weeks or longer is usually diabetic. 

An alimentary glycosuria occurs more readily if the sugar is eaten 
while the stomach is empty. Hunger lowers the assimilation limit con- 
siderably ; pregnancy also does the same. Diseases which may lower the 
assimilation limit are : cirrhosis of the liver, cerebral disease, poor nutrition,. 
fatty liver, phosphorus poisoning, the infectious diseases, certain neuroses, 
exophthalmic goiter, and any condition causing diuresis. Among the 
diseases which raise the assimilation limit are myxedema and pituitary 
gland disturbances. While most normal persons have an assimilation 
limit for glucose of 200 gms. or over, yet this varies much and 100 gms. 
has been accepted as the lowest limit of normal. 

The following method of determining the assimilation limit was recom- 
mended by Naunyn. Two hours after a breakfast consisting of coffee with 
milk (about 250 c.c.) and 80 to 100 gms. of bread, the patient ingests 100 
gms. of dextrose. If a definite glycosuria results the assimilation limit of 
that patient is pathologically lowered, while if the glycosuria is 1 % or over 
one has very good reasons for suspecting diabetes mellitus. The glycosuria 
begins in about 1 hour after the meal of glucose, reaches its maximum in 
from 2 to 4 hours and lasts at the longest but 8 to 10 hours. The assimila- 
tion limit for cane-sugar is practically the same as that for glucose (from 
150 to 200 gms.). Some prefer to use this sugar rather than glucose. 

The reasons why the breakfast precedes the test meal is that if the latter 
is received into an empty stomach the sugar may pass too rapidly to the 
bowel and escape digestion, and that some which reaches the lower part 
of the small intestine may be absorbed by the lymphatics rather than by 
the portal vessels and so not pass through the liver but at once enter the 
systemic circulation and so be excreted. 

That all glycosuria is not diabetic or that the prognosis of diabetes is 
not as dark as is usually believed is indicated by the observations of 
Barringer and Roper 52 who examined by repeated assimilation tests the 
condition of a series of patients in whom 5 years before a transitory spon- 
taneous glycosuria had been discovered. Of these, 20% had during this 
interval become definitely diabetic; 15% had probably, but not certainly, 
become diabetic; in the case of 10% there was much doubt as to whether 
their condition was diabetic or not; while 55% were surely not diabetic. 
These writers, therefore, refuse to agree with Von Noorden that all persons 
who show even occasionally a spontaneous glycosuria are of necessity cases 
of latent diabetes. 

To determine whether a transitory spontaneous glycosuria is diabetic 
or non-diabetic the patient's assimilation limit is tested a few months 
after the glycosuria has disappeared and again a f ew months later. \ 

52 Am. J. Med. Sc, June, 1907. 



164 CLINICAL DIAGNOSIS 

The improved methods of determining blood-sugar have recently made 
it possible to measure carbohydrate tolerance by the study of the blood 
rather than of the urine after a test meal (see page 552). This has 1 definite 
advantage since the variable threshold limit for sugar is eliminated. 

The Hunger Diabetes of Hofmeister. — Hofmeister found that if dogs 
under close confinement are kept on a poor diet, yet not starved, a certain 
number of them soon became diabetic and excrete 30% of the starch of 
their food as sugar. Naunyn believed that this is probably the explanation 
of some cases of that glycosuria in men which develop in cachexia producing 
diseases. Later Hoppe-Seyler reported 53 10 cases of temporary glycosuria 
in vagabonds whose lives had been unhygienic and whose diet had been 
miserable. Their glycosuria disappeared in 24 hours after their physical 
condition had improved somewhat. 

Glycosuria. — That a trace of glucose is present in normal urine can be 
proved by isolating glucosozone from large amounts of urine. Quantitative 
reduction tests before and after the fermentation of a normal urine also 
indicate the presence of this body. That is, a glycosuria is both normal 
and constant. 

When the output of glucose in the urine is sufficient in amount that the 
glucose may be detected by the tests in common clinical use the condition 
is termed a pathological glycosuria or a glycuresis. Theoretically, a 
glycuresis will develop: (1) When there is a hyperglycemia of about 
0.3% or over (see page 550). Such a hyperglycemia may be due to the 
ingestion of more sugar than can be warehoused or to the accumulation 
in the blood of glucose which the body cannot use and which therefore the 
kidneys will excrete. (2) When the ability of the kindeys to retain glucose 
is diminished, e.g., after the injection of phlorizin and in the interesting 
condition called renal diabetes. (3) When the glucose exists in the blood 
in some chemical combination which renders it unfit for use. 

A diabetic glycosuria differs from other glycosurias, according to Allen, 
in one fundamental particular: In the normal individual the greater the 
amount of sugar ingested the more is used, while in the diabetic individual 
the reverse is true (Allen's paradoxical law). That is, the limit of toler- 
ance in the diabetic individual is real, not apparent as in the normal indi- 
vidual, and if these limits are exceeded by the ingestion of more sugar 
than the diabetic individual can handle, his tolerance is at once lessened 
rather than increased as in the case of the normal individual. According to 
Allen, the diabetic condition is one in which there is a deficiency of pan- 
creatic amboceptor, by which term he designates a substance furnished 
by the pancreas which unites with glucose and forms a combination which 
the tissues can use. Glucose not in this combination circulates as a crystal- 
loid wh ; ch cannot be used by the cells, which acts as a diuretic and which 
is eliminated unchanged. In his earlier work Allen considered the problem 
* 53 Munch, med. Wochenschr., April-, 1900. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 165 

of diabetic glycosuria as essentially one of functional pancreatic fatigue 
and indeed in many ways this would describe the condition well. 
Clinically, cases of glycosuria may be classified as follows : 
i. The group of diabetic patients who have a definite limit to their 
glucose tolerance. In these cases one assumes always a disease of the pan- 
creas. In dogs whose pancreas has been destroyed or removed the glyco- 
suria may reach 10 or 22% and the animal lives not over 4 to 5 weeks. In 
these dogs practically all the sugar ingested is excreted in the urine and in 
an amount which bears a quite constant ratio to the urine nitrogen (2.8 : 1) . 
In no cases in man is quite all the glucose excreted and all patients 
have some tolerance to sugar. In some patients is found atrophy of the 
pancreas as a whole or, as in Opie's case, a degeneration limited to the 
islands of Langerhans. A very good illustration is reported by Mosenthal. 54 

2 . Glycosuria may follow oxygen starvation of the tissues due to many 
causes; suffocation and the death agony; certain poisons, as CO, curare 
and amyl nitrite; narcotics, as ether or chloroform. 55 The work of King, 
Mayle and Haupt, 56 however, throws much doubt on the belief that the 
glycosuria of ether anesthesia is due to oxygen starvation. 

3. Certain poisons, including morphia, strychnine, cocaine, 57 fusel oil, 
HgCl 2 , and certain acids can cause glycosuria. In this connection the 
work of Herter 58 is interesting. He showed that the local application to 
the pancreas of reducing substances (adrenal extract and various poisons, 
H 2 S, KCN, H2SO4, etc.) may cause glycosuria. 

4. Severe cooling of the body. 

5. The use of caffeine, theobromine, or any diuretic which stimulates 
the kidney may cause glycosuria (renal diabetes). This and phloridzin 
diabetes are the only cases of glycosuria which are not associated with 
hyperglycemia. Some patients with chronic nephritis have diabetes also, 
but, as a rule, the diabetes was their primary trouble. There is a tendency 
for the glycosuria of a case of nephritis to lessen because of an increased 
sugar tolerance as the nephritis progresses. This explains an old belief 
that B right's disease cures diabetes. 

Glycosuria sometimes follows a transfusion with normal salt solution, 
the injection of sugar into the blood, insults and injuries to the liver (well 
seen in animal experiments) , cirrhosis of the liver and diseases and injuries 
of the central nervous system, the best illustration of which in animals is 
the piqure of Claude and Bernard which causes an hyperglycemia of even 
0.7%, which may last from 6 to 48 hours, and a glycosuria which in rabbits 
may reach even 6%. In man a transitory glycosuria which may be some- 
what similar in nature sometimes follows apoplexy. This begins, as a rule, 

54 Arch. Int. Med., March 15, 1912. 

55 See also Brown, Johns Hopkins Hosp. Bull., May, 1900. 

66 Jour, of Exp. Med., August 1, 1912, vol. xvi, p. 178. 

67 See also Neubauer and Vogel, p. 92. 

68 Am. Med., 1902, p. 771. 



166 CLINICAL DIAGNOSIS 

about 2 hours after the hemorrhage, lasts even 6 days, and may reach even 
i or 2%. Glycosuria may also be associated with brain tumors, especially 
those of the base, often with dementia paralytica, epidemic cerebrospinal 
meningitis, tabes, multiple sclerosis, diseases of the sympathetic nervous 
system and severe trauma of the skull. This last group is interesting. The 
glycosuria may begin at once or even a year later and usually is perma- 
nent. Such cases as a rule are mild. In some one may suspect an interest- 
ing relation of diabetes insipidus since they begin as an intense sugar-free 
polyuria and later develop a glycosuria. Glycosuria is met with also in some 
patients with functional (?) neuroses, psychical disorders, in exophthalmic 
goiter, gout, arteriosclerosis and obesity. 

In 90% of the severe grades of hyperthyroidism there is present an 
hyperglycemia and in almost as many cases a glycosuria, either spontaneous 
or alimentary. In the milder cases an alimentary glycosuria and hyper- 
glycemia can usually be easily produced (2 hours after 100 gms. of glucose) 
and the return to fasting blood-sugar conditions is slower than normal. 

A hyperglycemia and often a spontaneous glycosuria can be obtained 
in hyperthyroidism following the subcutaneous injection of adrenalin 
(Goetch's test, see page 552). 

Emotional glycosuria. Folin, Denis and Smillie, 59 following the work 
of Cannon, Sbohl and Wright on animals, found glucose in the urine of 
22 of 192 patients suffering especially from depression, apprehension or 
excitement, and in that of 58 of 664 patients at another hospital for the 
insane. They then examined students just before and after hard examina- 
tions and found after the examinations sugar in the urine of 18% of the 
men and of 17% of the women, all of whom had been sugar-free before the 
mental strain. They therefore consider that pronounced mental and emo- 
tional strain may produce temporary glycosuria in man. 

Qualitative Tests for Glucose. — Since the most popular tests for 
glucose make use of copper solutions the students should study carefully 
the several reactions involved in the reduction of copper by a urine con- 
taining glucose, and, whatever the copper test he may later choose as his 
routine method he should first study Trommer's test, since in this each 
step in the reaction is taken separately and the chances of error are all 
very apparent. 

Trommels Test.— To a test-tube half full of urine is added about )i its volume of 10% 
NaOH or KOH and then a 10% solution of CuSCU in drops, until a few flakes of the 
Cu(OH) 2 which form do not disappear on slight shaking. The upper layer of the urine 
is then warmed. If sugar is present in pathological amounts a precipitate, yellow or 
red in color, appears at once at the top. The heating should then be stopped. The 
reduction of theeupric salt and the precipitation of the cuprous salts will spread through- 
out the fluid from above downward. The urine should always be examined while 
fresh. If much albumin is present this should be removed. 

69 Jour, of Biol. Chem., 1914, xvii, p. 519. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 167 

The reactions in the reduction of copper are as follows: If instead of urine pure 
water is used and the solutions of KOH and then of CUSO4 be added, the first drop of 
the latter will precipitate Cu(OH) 2 [CuS0 4 +2NaOH = Na 2 S0 4 +Cu(OH) 2 ]. The blue 
flakes of Cu (OH) 2 will blacken when heated since they will be changed to Cu(OH) 2 2CuO. 
At this point it should be emphasized that only a salt in solution can be reduced. If at 
the onset a little glycerin or some tartrate solution had been added to the water all of 
the Cu(OH) 2 formed would remain in solution and so would not blacken when heated. 
If, instead of glycerin or the tartrate solution, glucose had been added to the water, a 
similar blue solution of the Cu(OH) 2 would be obtained (due to CeHi2065Cu(OH) 2 ) but 
on warming this would be reduced and a yellow or red precipitate of cuprous salts 
would fall. 

In the normal urine are certain bodies which, like glycerin and the tartrates, will 
hold Cu(OH) 2 in'solution. Among these are the ammonia bodies, both those preformed 
and those resulting from boiling an alkaline urine, glucose and albumin if present. 
These, however, are not present in sufficient quantity to hold more than a drop or two 
of Cu(OH) 2 in solution and this trace when reduced will merely give a slight greenish 
color to the urine. If, however, to the normal urine or to the reagents used, glycerin, 
the tartrates, or an. excess of ammonia be added, more or all of the Cu(OH) 2 formed 
will be held in solution and this will assume an azure blue color which varies in depth 
with the amount of CUSO4 added. 

But the normal urine contains also reducing bodies other than glucose which will 
reduce the copper on warming. Among these are uric acid, the glycuronic acid com- 
pounds, pyrocatechin, bile pigments, creatinin and a trace of glucose, always present. 
The sum of all these reducing bodies would not make up a solution of over 0.5% if 
expressed in terms of glucose. These bodies, if present in normal amount, will reduce 
enough of the copper to give a dirty yellowish, not a clear yellow color to the solution, 
but not enough to give a precipitate. Sometimes there is in the urine an increased 
amount of these substances (other than glucose) to give a definite precipitate of cuprous 
salts; that is, a positive test may result. But uric acid does not reduce copper at a 
temperature of from 6o° to 70 C. as does glucose and creatinin reduces much copper 
only after long boiling, although it will a little at 6o° C. The presence of these and other 
bodies (uric acid, creatinin, ammonia, albumin) is, however, very important since cer- 
tain of them not only reduce copper but they all aid to hold in solution the small amount 
of the suboxides of copper which is always formed. Their ability in the normal urine 
to hold in solution these reduced suboxides is much greater than their ability to reduce 
copper and hence glucose may be added to normal urine even to a 0.5% solution before 
any cuprous oxide will precipitate. *- 

In the urine of a case of glycosuria there is a great increase of the glucose and, because 
of the polyuria, a decrease in the concentration of those bodies which prevent, the, pre^ 
cipitation of the cuprous salts, < -. 1 .-..■-:.• 

In performing Trommer's test it is important that the copper should not be added 
in excess since the black oxide formed by heat will mask the precipitate of the cuprous 
salts. In the case of normal urine from 3 to 5 drops of the CUSO4 solution>are sufficient 
to give a blue precipitate. In case sugar is present, however, one must continue: to/add 
the CUSO4 solution until the first flakes of Cu(OH) 2 remain undissolved. -For the .test 
to be positive a yellow (Cu(OH) 2 ) or red (Cu 2 0) precipitate must fall. 1 If- much sugar 
is present, metallic copper may be deposited on the glass (to clean such tesi->tube& strong 
nitric acid is recommended). In case there is less than 0.2% of sugar present mo Cuprous 
salt will precipitate, and yet the test may be very suggestive, because iofithe^clear bril- 
liant color of the yellow solution. Again, the precipitation of cuprous salt&> should 
occur at a temperature under the boiling point or promptly when the 'ruimeyis 5 just 
brought to that point to exclude the reduction by those bodies (creatinine urii;! amd; 
etc.) normally present. ■ . ,«■, <: >v» -a-I/o k> 



168 CLINICAL DIAGNOSIS 

For a successful test the proportions of the reagents should be accurately estimated. 
Since I part of sugar can reduce about 5 parts of Cu(OH) 2 , as nearly this amount of 
copper as possible should be present. Since glucose alone cannot dissolve as much of 
the Cu(OH) 2 as it could reduce were this in solution so glycerin, ammonia or the tar- 
trates are added to Fehling's, Purdy's, etc., reagents, to hold in solution as much Cu(OH) 2 
as possible in order that it may be at the disposal of the glucose. The optimum relation 
of reagents is 1 part of glucose to 5 (3 to 7) of Cu(OH) 2 and to 1 1 of NaOH. The excess 
of the last reagent is necessary since directly upon this does the temperature of reduction 
depend. If much too little NaOH be present, hours of boiling may be required to reduce 
the copper; if but 2 parts of NaOH to 1 of glucose are present a few minutes' boiling is 
enough, while if there is an excess of NaOH it is not even necessary to raise the solution 
to the boiling point in order to get a reduction. 

Again, the best chance of a precipitation of the cuprous salts obtains when there is 
in the urine a minimal amount of those bodies which would hold the reduced copper 
salt in solution. For this reason it is advised by many, as a matter of routine, to dilute 
the urine about 1 : 5 with water, since this dilution will reduce the influence of these 
bodies in much greater proportion than it will the reducing power of glucose. 

If to a urine rich in glucose considerable strong NaOH or KOH be added and only 
a little copper heat will produce a yellow, yellowish-brown or a dark-brown solution 
depending on the relative amount of sugar and alkali present (Moore's test). This 
dark color is due to the reaction of the alkali with that glucose in excess of the copper. 

The result of Trommer's test in a normal urine may be a clear yellowish solution 
or a grayish-green shimmer due to a slight precipitate of the copper compounds of the 
xanthin bases and uric acid. The copper precipitated by sugar is crystalline, while 
that by the xanthin bases is amorphous. To be positive for glucose the precipitation 
should take place while the urine is still hot and not after it has cooled down. If but a 
trace of sugar is present the precipitate may fall after long boiling or after cooling, but 
such a reaction is not positive. A brilliant yellow color of the clear urine after boiling 
may suggest the presence of sugar but does not prove it. In such a case if the test is 
repeated after the urine is diluted a characteristic precipitate may fall. Since a normal 
urine reduces some copper, and would reduce more could it hold more of the Cu(OH) 2 in 
solution, a normal urine which has become ammoniacal may give a positive test for sugar 
since the ammonia will dissolve the cuprous salt. In a normal urine it is possible some- 
times to get a positive test by adding an excess of NaOH and too much copper sulphate. 

The phosphate precipitate always present may be stained slightly yellow by the 
Cu 2 (OH) 2 formed even in normal urine. This often deceives. 

In all copper tests a considerable amount of albumin would not hinder the reduction 
by glucose, but would the precipitation of the cuprous salt and hence should be removed 
unless but a trace is present, in which case it may be disregarded. Among the reducing 
substances present in the urine which may give a positive test for glucose are allantoin, 
mucin, pyrocatechin, hydrochinon, urobilin and perhaps also indican. 

The test for glucose may be obtained when the glycuronic acid compounds are present 
in increased amounts, which is true following the use of chloral hydrate, chloroform, 
morphine, camphor, phenol, resorcin, thymol, and menthol. A positive reduction is 
obtained sometimes after the use of salicylic acid, benzoic acid, chrysophanic acid, 
oxalic acid, salol, thallin, santonin, copaiba, rhubarb, sulphonal, chloroform, acetphene- 
tidin, glycerin and after poisoning with KOH, H 2 S04, and arsenic. In alkaptonuria 
the test is positive. Saccharin hinders the reduction. 

This is not as delicate a test as is Fehling's or Benedict's, and yet we have been 
interested to observe that those who have had the greatest experience in sugar work 
use Trommer's as a routine test since it gives much more information about the urine 
than the mere presence or absence of glucose. From it one may get a hint of the presence 
of other reducing bodies also. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 169 

Fehling's Test Solution. — Fehling's solution contains Rochelle salts 
which aids to hold a maximum amount of the cupric hydroxide in solution 
and so allows a maximum formation of the cuprous salts. Fehling's solu- 
tion is made from 2 fluids, each quite permanent, but which must be kept 
separate, since an old mixed fluid may reduce on boiling. 

Solution A Solution B 

Copper sulphate, 34.65 gm. Rochelle salts, 173 gm. 

Distilled water, q.s. ad 500 c.c. Sodium hydrate, 125 gm. 

Distilled water, q.s. ad 500 c.c. 

Equal volumes of these 2 fluids are mixed and brought to a boil. As 
soon as one is certain that the fluid will not reduce itself the urine is added 
to the boiling reagent slowly until precipitation occurs or the amount of 
urine added equals % that of the mixed reagent used (never more). If 
then there is no precipitate the test is negative for glucose. The mixture 
should be brought again to the boiling point after each addition of urine 
but prolonged boiling should be avoided. A precipitate which forms after 
the urine has been allowed to stand does not indicate sugar. By adding 
the urine slowly one can estimate pretty accurately the amount of sugar 
present. This test is positive for 0.08% of glucose. Although more deli- 
cate, it should be remembered that this test has all the faults of Trommer's. 
A normal urine will always reduce a little copper, but not if the urine is 
first diluted so that its specific gravity is about 1.005 (Zeehuisen). 

Benedict's Test. — The following directions for making Benedict's 
reagent should be carefully followed. 

Gms. or c.c. 

Copper sulphate (pure crystallized) 17.3 

Sodium or potassium citrate I73-Q 

Sodium carbonate (crystallized) (one-half the weight of the 

anhydrous salt may be used) 200.0 

Distilled water to make 1000.0 

The citrate and carbonate are dissolved together (with the aid of heat) 
in about 700 c.c. of water. The mixture is then poured (through a filter 
if necessary) into a large beaker or casserole. The copper sulphate (which 
should be dissolved separately in about 100 c.c. of water) is then poured 
slowly into the first solution, with constant stirring. The mixture is then 
cooled and diluted to 1 liter. This solution keeps indefinitely. 

Five cubic centimeters, a trifle over 1 teaspoonful, of the Benedict 
solution are placed in a test-tube and 8 to 10 drops (not more) of the urine 
to be examined are added. By using always these exact quantities very 
delicate results are obtained. The mixture is then heated to vigorous 
boiling, kept at this temperature for 3 minutes, and allowed to cool spon- 
taneously. In the presence of glucose the entire body of the solution will 
be filled with precipitate, which may be greenish, yellow or red in tinge, 



170 CLINICAL DIAGNOSIS 

according to whether the amount of sugar is slight or considerable. If 
the quantity of glucose be low (under 0.3%) the precipitate will form 
only on cooling. If no sugar is present the solution either remains perfectly 
clear, or shows a faint turbidity that is blue in color and consists of pre- 
cipitated urates. Benedict states that the test if performed as above will 
detect glucose in as low concentration as 0.01 to 0.02% provided the urine 
is of low dilution. 

Almen-N y lander' s Test. — The Almen-Nylander solution is made up by 
dissolving 4 gms. of Rochelle salt in 100 c.c. of 10% NaOH (sp. gr. 1.015) 
warming, and saturating this solution with bismuth subnitrate (about 
2 gms. are necessary). The solution is then cooled and filtered and kept 
in a dark bottle. It is permanent for years. 

To the urine in a test-tube is added Ko its volume of this reagent. The 
fluid is then boiled preferably in a water bath for 5 minutes. If sugar is 
present the urine will turn black and a black precipitate of metallic bismuth 
will be deposited. If the urine contains over 0.2% of glucose the yellow 
color of the Moore test is first seen. Should the specimen become black 
only after it has been cooled the test is not necessarily positive. If but a 
trace of glucose is present, the white sediment of phosphates may be only 
slightly gray, especially on its upper surface. The boiling should be con- 
tinued for fully 5 minutes, since only too often will the urine suddenly 
darken after one is tempted to pronounce it negative. If but a trace of 
sugar be present, the amount of urine and of reagent used be accurately 
measured. If this test is negative we may be sure that no sugar is present. 
If it is faintly positive, the result should be confirmed, since bismuth is 
reduced also by the quantity of paired glycuronic acid compounds some- 
times present. This test is very delicate. It will indicate 0.05% (others 
say 0.025%) of glucose. Some claim that it is positive in 14% of normal 
urines especially if these are concentrated. Uroerythrin and hematopor- 
phyrin both may give a similar test. Any modification in the reaction of 
the mixed reagent and urine injures the delicacy of the test, hence it should 
be applied carefully if the urine is ammoniacal since the NaOH of the 
reagent will replace the ammonia, which will be lost by volatilization 
leaving the solution not alkaline enough. Rhubarb and senna will reduce 
the bismuth salt but the fluid before heating will take a brownish-red 
color. This test is positive after salol, benzol, sulphonal, trional, antipyrin, 
kairin, much quinine, eucalyptus tincture and oil of turpentine. It is 
also positive after a person has eaten asparagus, a fruitful source of error. 
All albumin should be removed unless but a mere trace is present, since 
the Bi 2 S 3 , usually reddish in color, will, if precipitated in considerable 
amount, look black. This test is very valuable as a control of the copper 
tests, since this bismuth reagent is not reduced by uric acid, creatinin, 
pyrocatechin, hydrochinon, nor the alkaptdn bodies, and these are the 
greatest sources of error if copper solutions' &re used. fcfcj <f» 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 171 

Fermentation. — The fermentation test is one of the best for the detec- 
tion of glucose. If a busy clinician used this only fewer mistakes would 
be made. Only those sugars which have 3 , or a multiple of 3 , carbon atoms 
(and not all of these) will ferment with gas production. A piece of fresh, 
active yeast about the size of a pea is mixed with the urine in a beaker 
by rubbing it against the side of this vessel with a glass rod until the lump 
is well broken up. The urine should not be shaken during this process 
since enough air will be taken up later to give a bubble suspiciously large. 
The urine is then transferred to a fermentation tube and let stand for a 
few hours at a temperature of from 15 to 34 C. Two control tests should 
always be made : the 1 with normal urine to which the yeast and a little 
glucose have been added to prove the activity of the yeast; another fer- 
mentation tube is filled with normal urine plus a little of the same yeast 
to rule out the possibility that the yeast will undergo self -fermentation. 
All fermentation stops at 45 C. and over. If sugar is present the rapidity 
of gas production will depend to a certain extent on the amount of yeast 
added. The amount of gas formed from a given amount of sugar will, 
however, depend in part on the age of the yeast used since less will be 
formed by an old yeast. The maximum production of C0 2 (this would 
mean that 46.5% of the sugar was thus split) is obtained when for each 
gram of sugar is added not more than % a gram of fresh yeast. If too much 
yeast is used self -fermentation may result. This test if applied to a pre- 
viously boiled urine will indicate from 0.1 to 0.05% of glucose. 

Some consider it necessary to prove that the gas which collects in the 
fermentation tube is C0 2 . This is easily done by dissolving it in NaOH. 
Some consider it necessary to prove that alcohol also is formed in this 
fermentation. This is easily done by distilling the fermented urine, adding 
to the distillate a little NaOH and some Lugol's solution, then warming 
and allowing the fluid to stand for several hours. Crystals of iodoform 
will appear if alcohol or acetone was present in the distillate. Or, to the 
distillate may be added a little very dilute solution of potassium bichromate 
and a little sulphuric acid. This fluid when heated will turn green and 
give off the odor of aldehyde. > 

Some bacteria will produce gas if the urine is allowed to stand too long. 
For that reason the acitivity of the yeast should be judged at the end of 
not over 6 hours. Or, bacterial growth may be inhibited by the addition 
of enough NaF to make a 1% solution, or by tartaric acid. Many recom^ 
mend that the urine to be examined first be boiled for about 10 minutes 
to sterilize it and also to free it from air. 

If but a trace of glucose is present in the urine no bubble of C0 2 may 
appear in the fermentation tube since urine can hold some of this gas in solu- 
tion. For this reason a urine may be negative for glucose by the fermenta- 
tion test and yet positive to Nylander's test, in which case the latter test 
should be repeated using the fermented urine which then will be negative. 



172 CLINICAL DIAGNOSIS 

Phenylkydrazin. — The phenylhydrazin test is theoretically the court of 
last appeal in the recognition of those carbohydrates which form with this 
reagent osazones of definite crystalline shapes and definite melting points. 
Albumin must be removed from the urine to be tested for it may hinder 
crystallization. Of the many methods of applying this test clinically that 
of Cipollina 60 is the best. In a common test-tube are mixed 5 drops of 
pure phenylhydrazin (the base), 0.5 c.c. of glacial acetic acid, and 4 c.c. 
of urine. This mixture, constantly shaken to prevent sputtering, is boiled 
over a low, free flame for about 1 minute, at the end of which time 4 or 5 
drops of NaOH solution (sp. gr. 1.16) are added to reduce the acidity, 
although the fluid must still remain acid after the addition of the alkali. 
The liquid is then boiled again for a moment and cooled. The character- 
istic rosettes of crystals will form at once or at least within 20 minutes. 
The best results are obtained with urines of low specific gravity. 

If much glucose is present the crystals of phenylglucosazon will form 
a yellow crystalline deposit in the tube. This may be filtered out, dis- 
solved in hot 60% alcohol and recrystallized by adding water and boiling 
the alcohol away. The melting point of the crystals should then be deter- 
mined. If pure, this lies between 204 and 205 C. ; when impure, from 
1 73 to 194 C. These crystals are yellow needles arranged in sheaves, 
which are difficultly soluble in water and in hot absolute alcohol, are easily 
soluble in 60% hot alcohol and which will crystallize out if water be added 
and the alcohol evaporated off. They are insoluble in ether, chloroform, 
etc., but are soluble in glacial acetic acid. Their solution is levorotatory. 

A very simple method of determining the melting point of crystals 
is as follows (see Fig. 30) : A small flask, A , is filled % full with concentrated 
sulphuric acid. Through a perforated stopper is inserted a test-tube, B, 
also % full of the same acid. Into this dips a thermometer, C, to 
which is attached a tube, D, with a lumen about 1 mm. in diameter and 
sealed at the lower end which contains the dry crystals to be tested. This 
tube is attached by a rubber band to the thermometer. The flask is 
warmed slowly with a Bunsen burner and the point noted at which the 
crystals melt. 

For other forms of apparatus the reaaer is referred to Menge's report. 61 

This test theoretically is very delicate and in simple solutions of pure 
glucose should show 0.003% of the sugar. Not all of the glucose is pre- 
cipitated, the amount depending on the concentration of the glucose and 
the relative proportions of the reagents used. From a 5% pure glucose 
solution the maximum precipitation obtained by Fischer was from 85 to 
90%. Much depends on the purity of the phenylhydrazin. In practical 
urine examinations this test is not delicate and seldom if ever indicates 
glucose in urines which will not reduce Fehling's solution. 

60 Deut. med. Wchschr., 1901, Not. 21, p. 334. 

61 Bull. 70, Hygienic Lab., U. S. A., Oct., 1910. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 173 



The greatest danger of error when using this test is from pentose, but 
these crystals will melt at 159 to 160 C. The glycuronic acid compounds 





Fig. 30. — Melting point of crystals. A, flask, and B, 
test-tube of sulphuric acid; C, thermometer; D, fine- 
bore tube for crystals. 

will form similar crystals but their melting point is lower, 114 to 115 C. 
All of the sugars which will reduce copper will give crystals, especially 
pentose and lactose. 



174 



CLINICAL DIAGNOSIS 



To form these crystals with lactose one must concentrate the urine 
and extract the lactose. Those sugars which differ only in the first 2 carbon 
atoms, the rest of the graphic formula the same, as, for instance, glucose, 
fructose, and mannose, all give exactly the same osozone. Crystals would 
be formed with acetone, hydrazin, oxalic acid, and uric acid, but in human 
urine not in sufficient amount to lead to error. 

Zunz 62 considers the phenylhydrazin test one of great clinical value 

Together with the crystals one finds also in the precipitate brown 
scales and oily droplets. These form even when a pure solution of glucose 
is used. This by-product is C12H12N2, and" can be removed by washing 
the sediment with chloroform or 95% alcohol and then recrystallizing the 
glucosazone from 60% alcohol. Sometimes even in the presence of glucose 
in the urine one gets a precipitate consisting only of brown scales or yellow 
amorphous granules or droplets. Such a result is to be considered negative, 
and yet glucose may be present. 

In review it may be emphasized: that there is nothing characteristic 
in the shape of these crystals ; their best solvent is 60% alcohol ; they are 
best recrystallized by pouring the 60% alcoholic solution into water and 
evaporating off the alcohol; the hot glacial acetic acid solution, which is 
soon destroyed, may be tested with the polariscope; that in testing the 
melting point one cannot expect to find it exactly 2 04 to 205 C, so much 
depends on the purity of the crystals and on the speed with which the tem- 
perature is raised. Glucosazon differs from galacosazon in that its glacial 
acetic acid solution is levorotatory while that of the latter is optically 
inactive; otherwise they are very similar. 

For the further separation of the carbohydrates of the urine we copy 
the table of Zunz. 

Fermentation f Dextrorotatory, 
positive. { Levorotatory . 

Fermentation 
negative 

Give orcin reaction. 



Gives crystals 

with 

phenylhydrazin 

directly in 

urine. 



Melting point 

of crystals 
about 200 C. 



Glucose. 
Levulose. 

Lactose. 



Melting point 

of crystals 
about 150 C. 



Pentoses. 



Isomaitose. 



Do not give orcin reaction. 

Gives crystals with phenylhydrazin only after the urine has I Paired' glycuronic 

been warmed with dilute sulphuric acid. acid compounds. 

The polariscope is a clinical instrument of great value in the recognition 
of sugars. It should be borne in mind that most normal urines are slightly 
levorotatory and that some urines which contain no dextrose are dextro- 
rotatory (e.g., Borntrager, in 2 morphia habitues) and so traces of sugar 
may escape detection or be wrongly reported as present. The ordinary 
instrument will detect 0.2% or more of glucose. The test is of greatest 
value if the urine be examined before and after fermen tation 

02 Jour. Med. de Bruxelles, July 10, 1902. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 175 

Heat Test. — An easy test for glucose, sometimes valuable, more delicate than one 
would imagine, and always possible, is the following: one drop of urine is evaporated to 
dryness in a porcelain dish. It is then warmed gently. The residue becomes yellowish 
brown and at a temperature of 190 to 200 C. gives off the odor of caramels. 

Moore's Test. — Moore's test was one of the first used for sugar. Its value now is 
not so much as a test but as a reaction which is often met with unexpectedly and should 
be correctly interpreted. To the urine is added % volume of a strong solution of KOH 
or NaOH. On warming there results, first, a yellow, then an orange, and finally a dark 
brown color with an odor of caramels, more distinct if the urine is acidified. It may 
be necessary to boil for some time. This reaction develops slowly at room temperature. 
It is not a delicate test nor very accurate, since some normal urines will darken somewhat, 
also urines which are rich in mucus. The names glucinic acid and melasinic acid have 
been suggested for the substance coloring the urine. 

Choice of Method. — If any of the above-described reduction tests is 
definitely positive, sugar is present. Of these, Nylander's is the one to be 
recommended first. If it is negative then no sugar is present. If positive, 
fermentation should next be tried. If this is positive the sugar probably 
is glucose, but levulose must be excluded. For the practitioner the fer- 
mentation test is perhaps the best test, since it leads to least confusion. 

If one desires to clear the urine of glucose for further tests it must be 
remembered that glucose is not precipitated by sugar of lead but is almost 
completely by basic lead acetate. 

Quantitative Determination op Glucose. — When sugar is known 
to be present in a specimen of urine and in good amount, an approximate 
quantitative estimate is possible by the use of Naunyn's table : 

2 liters of urine of specific gravity 1028 to 1030 = 2 to 3% 

3 liters of urine of specific gravity 1028 to 1032 =3 to 5% 

5 liters of urine of specific gravity 1030 to 1035 =5 to 7% 

6 to 10 liters of urine of specific gravity 1030 to 1042 =6 to 10% 

In estimating the amount of sugar from the specific gravity of the urine, 
and in using the following formula which employs the coefficient 230, it is 
assumed that changes in the specific gravity of urine are due to but 1 vari- 
able, i.e., sugar. This is not strictly true since urea and the chlorides also 
are variables. 

Suppose a diabetic patient voids 3 liters of urine in 24 hours and that 

this has a specific gravity 1.030. A normal person will void approximately 

2 liters in 24 hours and the specific gravity would be 1.015. To determine 

the specific gravity of this normal person's urine were he to void 3 liters, 

we would calculate as follows : 

2X1. 015 + 1.000 

. = 1.010 

3 
1.030 — 1. 010 = .020 
.020X230=4.6 

That is, the urine of the above-mentioned diabetic patient would con- 
tain approximately 4.6% of glucose. 



176 CLINICAL DIAGNOSIS 

Supposing the patient voided in 24 hours 6 liters of urine with a specific 
gravity of 1.030. 

2X..oi5+4.ooo = I005 

1 .030 — 1 .005 = 0.025 
0.025X230 = 5.8% 

Benedict's Quantitative Test. — Benedict's method is a great improve- 
ment over Fehling's and its several modifications. In Benedict's, as in 
Fehling's, method the glucose of the urine reduces a definite amount of 
copper in alkaline solution, thus decolorizing to a quantitative degree the 
blue copper solution. In Benedict's method, however, the copper is precipi- 
tated as cuprous sulphocyanate, a snow-white compound, and the end reac- 
tion is therefore much sharper than in Fehling's test. The solution for 
quantitative work, which keeps indefinitely, has the following composition : 

Pure crystallized copper sulphate , 18 gms. 

Crystallized sodium carbonate (or 100 gms. of the anhy- 
drous salt) 200 gms. 

Sodium or potassium citrate 200 gms. 

Potassium sulphocyanide 125 gms. 

Five per cent, potassium ferrocyanide solution 5 c.c. 

Distilled water to make a total volume of 1000 c.c. 

With the aid of heat dissolve the carbonate, citrate, and sulphocyanide 
in enough water to make about 800 c.c. of the mixture and filter if necessary. 
Dissolve the copper sulphate separately in about 100 c.c. of water and 
pour the solution into the other liquid, with constant stirring. Add the 
ferrocyanide solution, cool and dilute to exactly 1 liter. Of the various 
constituents the copper salt only need be weighed with exactness. Twenty- 
five cubic centimeters of the reagent are reduced by 50 mg. (0.050 gm.) 
of glucose. 

The procedure for the estimation is as follows: The urine, 10 c.c. of 
which should be diluted with water to 100 c.c. (unless the sugar content 
is believed to be low), is poured into a 50 c.c. buret up to the zero mark. 
Twenty-five cubic centimeters of the reagent are measured with a pipet 
into a porcelain evaporating dish (10 to 15 cm. in diameter), 10 to 20 gms. 
of crystallized sodium carbonate (or % the weight of the anhydrous salt) 
are added together with a small quantity of powdered pumice stone or 
talcum and the mixture heated to boiling over a free flame until the car- 
bonate has entirely dissolved. The diluted urine is now run in from the 
buret, rather rapidly until a chalk- white precipitate forms and the blue 
color of the mixture begins to lessen perceptibly, then a few drops at a time 
until the disappearance of the last trace of blue color, which marks the 
end point. The solution must be kept vigorously boiling throughout the 
entire titration. 

If the mixture becomes too concentrated during the process, water may 
be added from time to time to replace the volume lost by evaporation; 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 177 

however, too much emphasis cannot be placed upon the fact that the solu- 
tion should never before or during the process be diluted to more than the 
original 25 c.c. Moreover, it will be found that in titrating concentrated 
urines, or urines with small amounts of sugar, a muddy brown or greenish 
color appears and obscures the end point entirely. Should this be the 
case the addition of about 10 gms. of calcium carbonate does away with 
this difficulty. 

The calculation of the percentage of sugar in the original sample of 
urine is very simple. The 25 c.c. of copper solution are reduced by exactly 
0.050 gm. of glucose. Therefore the volume of diluted urine drawn out 
of the buret contains 50 mgms. of sugar. 

When the urine is diluted 1 to 10, as in the usual titration of diabetic 
urines, the formula for calculating the fjercentage of sugar is the following: 

^^- X 1 000 = percentage in the original sample, wherein X is the number 

of cubic centimeters of the diluted urine required to reduce 25 c.c. of the 
copper solution. 

Chloroform should not be present in the urine during the titration. 
If it had been used as a preservative it may be removed by boiling a sample 
for a few minutes and then diluting to the original volume. 

To determine the glucose in a specimen of urine containing very little 
sugar, about 1500 c.c. of the urine is precipitated with sugar of lead and the 
nitrate then precipitated with basic lead acetate and a little ammonia. 
This precipitate is suspended in alcohol and decomposed with H 2 S. 
The filtrate is then cleared with animal charcoal if necessary and 
evaporated at low temperature to a small volume. The amount of glucose 
in this solution is then determined with the polariscope. A correction 
must be made for lactose or bile acid if present, both of which are dextro- 
rotatory. To do this the alcohol after the specimen is polarized is evapor- 
ated off, the residue dissolved in water, yeast added and the glucose removed 
by fermentation. This fluid is then filtered, the precipitate washed with 
alcohol, filtered, the filtrate restored to the original volume, and it is again 
polarized. From the difference between these two readings the amount of 
glucose may be calculated. 

Polariscope. — For this very important test and quantitative method 
the specimen must be very clear, so clear that even fine type may very 
easily be read through the tube of the polariscope full of the urine. All 
albumin must be removed since it is levorotatory. The urine is best 
cleared, if possible, with infusorial earth. An excess of this is added to 
the urine which then is well stirred and filtered. The filtrate should be 
poured back into the funnel until it runs clear. If, as sometimes happens, 
this method does not clear the urine perfectly, crystals of sugar of lead 
are to be added to the infusorial earth and the urine filtered through 
this mixture. 
12 



178 



CLINICAL DIAGNOSIS 



Sugar of lead in crystals is somewhat preferable to it in solution since 
the latter would change the volume of the urine. Yet some prefer to add 
to 90 c.c. of the urine 10 c.c. of a PbAc solution (25 gms. in 100 c.c.) and 
then filter. This clears the urine perhaps better. The proper correction 
for the change of volume is easily made. Basic lead acetate cannot be 
used. Infusorial earth is to be recommended since lead acetate in any 

E _^ 




Fig. 31. — Half-shadow saccharometer. A, ocular used in focusing the field; B, graduated disk; C, 
vernier; D, lever for rotating analyzer; E, tube for urine; F, glass disk; and G, cap for end. 

excess will alter the physical properties of the urine and will certainly 
remove some of the glucose, although it would none from a pure glucose 
solution. Yet infusorial earth also may remove some of the sugar. Others 
recommend that the urine be cleared with a small amount of PbAc and a 
teaspoonful of Na 2 S0 4 (added after the sugar of lead is dissolved). 

The tube of the polariscope is filled with the clear urine, care being 
taken that no air-bubbles be enclosed, and the angle of rotation measured 
by the scale on B (Fig. 31), using the vernier C, to read the fractions 
of a degree. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 179 

The tube, E, is first cleaned thoroughly and dried. The glass disks, 
F, are made perfectly clear. One glass is fastened in place and then the 
tube filled with the perfectly clear urine till the meniscus is convex. The 
second glass disk is then slid in position from the side, pushing off the 
excess of urine and leaving no air beneath. The metal cap, G, is then 
screwed down over this. 

The student should understand thoroughly the instrument that he is 
using. There is a great variety of polariscopes on the market which differ 
slightly in their construction and more in their accuracy. It is seldom, 
of course, that a standard polariscope is used. The most are instruments 
modified for clinical purposes. The polariscope is an instrument which 
measures the angle of rotation of polarized lights caused by an optically 
active substance; the length of the standard tube is 10 cm. or a multiple 
of this and the readings are made in degrees. The specific rotation of glu- 
cose is {a) D = 52.74°. If a polariscope is used, therefore, to examine urine 
the angle of rotation must be divided by 0.527 to give the percentage of 
sugar in the urine. The instruments in clinical use are usually " half- 
shadow instruments " whose tubes are of such length (188.6 mm. but 
better 189.4 mm.) that i° of rotation will indicate 1% of glucose. Each 
instrument usually has a second tube, ){ as long (94.3 mm., better 94.7 mm.) 
for highly colored urines. Another instrument which has become very 
popular and which is more convenient to use is the saccharometer in which 
the rotation by the sugar is balanced by a compensating quartz wedge 
on which is marked an empirical scale. The great advantage of this instru- 
ment is that an ordinary white light, as the Welsbach burner, can be used 
while in the other instrument a sodium flame alone is admissible. One 
disadvantage of this instrument for teaching purposes is that the principles 
in optics involved are not as evident to the student. 

In using the half -field polariscope the field must be first focused at A 
and the zero point determined. This changes somewhat with the tempera- 
ture, particularly in a carelessly used instrument. The tube is then inserted, 
the field again focused sharply, and the rotation determined. The accu- 
racy with which this can be done will depend upon the clearness of the 
field, which in turn will depend on the fluid, the focus, the sensitiveness of 
the instrument, and the brightness of the light. There are two methods in 
common use of finding the end-point at which the fields are of equal illumi- 
nation. According to one the analyzer is rotated until a black band seems 
to cross the division of the fields. This shadow, purely subjective, always 
appears a little too soon, therefore several readings from both directions 
should be made and averaged. According to the second method the analy- 
zer is slowly turned, always in the same direction, and using the eye but 
for a few seconds at a time, until the end-point seems to be just reached. 
This point also will be attained a little too soon, hence several readings 
should be made, turning the analyzer from both directions, and an average 




180 CLINICAL DIAGNOSIS 

taken. In all cases it should be remembered that the eye should be used 
for not over 1 5 seconds at a time to avoid fatigue of the retina. The depth 
of the illumination of the whole field should be judged and not of the con- 
tiguous portions of the 2 shadows (see Fig. 32). 

Many of these half -shadow instruments are so sensitive that they indi- 
cate a difference of even 0.02 , but since the error inherent in the urine 
itself is even 0.2% it is evident that very minute readings have only the 
appearance of accuracy. 

The surface of the ends of the tube must be planned accurately to a 
right angle to its long axis, otherwise the tube cannot be used. This errcr 
can easily be detected by putting the empty tube in the instrument, focus- 
ing carefully, and then revolving the tube around its long axis, which 
will produce the same effect as though the analyzer were rotated. Leather 
washers are necessary between the metal caps and the glass disks to prevent 

O 

Fig. 32. — The fields as seen in the two most common 
types of clinical saccharometers. The central figures, 
gray fields with halves of equal illumination, are the zero 
points. The others are the fields with too little and too 
much rotation. 

tension in the glass from the pressure of the cap, since glass subjected to 
too high tension becomes doubly refractive. For this reason, before any 
readings are made, the tube properly inserted in the instrument should 
be rotated around its long axis and the effect watched through the analyzer. 
If this rotation causes the 2 fields to change in relative intensity 1 of the 
above mentioned sources of error is present. Differences in the intensity 
of illumination of the whole field may indicate either that the solution is 
not homogeneous or that the tube is dirty. 

Normal urine is slightly levorotatory (0.005 to 0.18 ). A trace of sugar 
may therefore be present and cause no dextrorotation. In some cases 
the urine is dextrorotatory when glucose is not present. Such was true 
of 2 patients with the morphia habit (Borntrager) . 

On the whole this is the quickest practical qualitative method of sugar 
analysis. Albumin must be removed from the urine since it is levorotatory. 
Should the worker have occasion to make up a glucose solution as a control 
he must remember to use in the polariscope a, solution which has stood 
for at least 1 day, since glucose when first dissolved shows some birotation. 




THE URINE: CARBOHYDRATES AND ALLIED BODIES 181 

Fermentation. — According to one method (Robert's method) the amount 
of sugar in the urine is estimated from the difference in the specific gravity 
of the urine before and after fermentation. The urine should first be acidi- 
fied, if necessary, with tartaric acid. The specific gravity is then carefully 
determined, using a very accurate aerometer and making the proper cor- 
rection for the temperature of the fluid. A piece of washed yeast the size 
of a hazelnut is then added and the urine allowed to ferment at from 15 
to 35 C. (a temperature of 34 C. is the best) until it gives no qualitative 
test for 'sugar. This usually takes from 24 to 48 hours. The sediment is 
then brought into suspension and the specific gravity of the urine again 
tested. The difference between these specific gravities multiplied by 234 
will give the percentage of sugar. For this determination it is best to use 
the pycnometer method to determine specific gravity, since the aerometrical 
method, which at the best is poor, is hardly delicate enough for this work. 
If accurately applied the results are correct to about 0.1%. Albumin 
need not be removed. This method can be used if 0.5% or more of sugar 
is present. This is the best method for the practitioner who has not a 
polariscope nor the time to prepare for and to carry out the titration 
method. Yet unless one is careful in the details, an error of even 5% 
may be made. 

Fermentation: Gas-Volumetric Method. — The Einhorn method of esti- 
mating the amount of sugar in a specimen of urine by measuring the amount 
of carbon dioxide produced by fermenting it with yeast is much more accur- 
ate than is generally believed, provided one uses a fresh yeast and pays due 
attention to temperature, etc. Lohnstein's 63 instrument is said to be accur- 
ate and to give the final result in 6 hours. 

Levulose is a sugar which is met with widely in the vegetable kingdom, 
particularly among the fruits. It is often present in the urine of diabetics, 
but practically always in company with glucose. An alimentary levulo- 
suria can usually be produced by injecting or feeding levulose to those 
cases with liver trouble which seem to disturb the ability of this organ to 
transform levulose to glycogen (see page 182). In addition to these 2 forms 
of levulosuria there are on record a few, 6 or 7 only, cases of pure levulosuria 
(Naunyn) with even from 1 to 2% (as a rule, less than 1%) of this sugar 
in the urine. Rosin and Laband reported 64 an interesting case of pure 
levulosuria with about 0.6% of levulose in the urine. This patient showed 
a levulosemia of 0.5% even when the urine was negative. The levulosuria 
of this patient was uninfluenced by the ingestion of even 100 gms. of levu- 
lose or glucose. In a case reported by Strouse and Friedman 65 the levulose 
of the urine seemed to be entirely exogenous and derived from ingested 
levulose and from higher carbohydrates, which contained the levulose 

63 Munch, med. Wochenschr., 1899, No. 50. 

64 Zeitschr. f. klin. Med., 1902, vol. xlvii, p. 182. 

65 Arch. Int. Med., January 15, 1912, vol. ix, p. 99. 



182 CLINICAL DIAGNOSIS 

molecule. The levulosuria of this patient was not affected by the glucose 
ingested nor was his tolerance for glucose decreased. Levulose can be 
used easily by some diabetics, but by no means all. As a rule if levulose 
is ingested in large amounts almost all of it is excreted as glucose. 

The diagnosis of many of the reported cases of levulosuria is doubtful, 
since levorotatory bodies often present in the urine were not excluded. 
Levulosuria is to be suspected when the percentage of sugar as determined by 
polarization is definitely less than that which titration would indicate and 
when the levorotatory body is found to be fermentable with gas production. 

The most important levorotatory sugars are laiose and fructose. Fruc- 
tose gives reactions very similar to those of glucose ; it reduces copper some- 
what less readily (10%), it ferments with gas production and has an angle 
of levorotation of uncertain amount. Its characteristic test is that of 
Seliwanoff , although Rosin, also Fr. Miiller 66 state that glucosamin also 
will give this test. To a warmed solution of resorcin in moderately dilute 
hydrochloric acid (i volume of HC1 to 2 volumes of H 2 0) is added a little 
of the sugar in question. If it be levulose the fluid at once takes on a 
beautiful red color, due to a substance which precipitates on cooling and 
which is soluble in alcohol. Levulose, if warmed with a concentrated 
alcoholic solution of resorcin, gives a brick-red color. It gives the same 
osazone as glucose. It is a more fragile body than is glucose. 

The other levorotatory bodies of the urine which must be excluded are 
albumin, glycuronic acid compounds, /3-oxy butyric acid, and cystin. If 
the levorotation of a urine disappears on fermentation, the chances are 
that levulose was present. This should be confirmed, however, by isolating 
the sugar and testing it in pure solution. 67 

The alimentary levulosuria test for functional disturbance of the liver has 
attracted considerable attention. Strauss 68 found that the ingestion of 
100 gms. of levulose was followed by a levulosuria in 90% (26 of 28 cases) 
of the cases with hepatic trouble which he studied, and in but 11% (6 to 58) 
of the normal men. Ferrannini and B ruining considered the test valuable, 
but Landsberg 69 could obtain the test in but 9 of 2 1 cases of liver trouble 
(not severe ones), and in 4 of 7 normal persons. He therefore doubts that 
it has any value. 

Lactose is found in the urine of the great majority of women during 
lactation. Ney found it in 115 of 148 cases, others in all. The output 
reaches its maximum on the second to the fourth day after delivery. The 
amount eliminated usually is small, but it may reach even 2 to 3%. In 
these cases the sugar is absorbed from the lacteal glands. Lactose appears 
in the urine also of patients who for a long time have been on a milk diet. 
It will as a rule appear in the urine also after the ingestion of 100 gms. of 

66 Deutsches Arch. f. klin. Med., 1904, p. 1630. 

67 See Peligot method, Compt. rend., vol. xc, p. 153. 

68 Deutsch. med. Wochenschr., 1901, Nos. 44 and 45. 

69 Ibid., August 6, 1903. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 183 

this sugar. Voit found that if lactose be fed to diabetics they excrete more 
glucose, while in the case of lactating women this is not true. 

Lactose is dextrorotatory (52.5 ). Lactosazon crystallizes in small 
yellow prisms arranged in spheres whose melting point is 200 C. (This 
test cannot be applied to urine directly. The urine must be concentrated 
and the lactose extracted.) Its reduction tests are similar to those of glu- 
cose, but copper is reduced somewhat less actively and ammoniacal silver 
nitrate is reduced without the aid of heat. Nylander's test is positive. 
If a solution of lactose is boiled for several hours with dilute mineral acid, 
the lactose will be inverted to galactose and glucose. Lactose is not fer- 
mented by ordinary yeast. It can be fermented but without the production 
of CO 2 . The presence of lactose in the urine is to be suspected when the 
copper and bismuth tests are positive yet somewhat slow, and the fermenta- 
tion and phenylhydrazin tests are negative. Urine to be tested for lactose 
should first be sterilized, otherwise bacteria will split lactose giving rise 
to a fermentable sugar. If a urine does not ferment with gas production 
and yet does reduce copper, lactose or pentose should be suspected. 

Rubner's test for lactose deserves special mention. If urine containing 
lactose is boiled from 3 to 4 minutes with an excess of sugar of lead it 
becomes yellow or brown in color. While still hot, ammonia is added until 
the precipitate which forms no longer dissolves. From the intense brick- 
red fluid which results, a copper-red precipitate will separate, leaving the 
supernatant fluid colorless. If the specific gravity of the urine to be tested 
is over 1.020 it is wise to dilute it one-half. Glucose under these conditions 
would give a red solution with a yellow precipitate; maltose, a yellow 
solution, and levulose, no change of color at all. 

Pentoses. — The pentoses are a very important group of sugars with a 
chain of 5 carbon atoms. These are rather complex carbohydrates from 
which pentose may be split by hydrolysis are widely distributed in the 
vegetable kingdom. In the metabolism of the herbivora the pentoses play 
almost the same role as the hexoses in man, in that they are glycogen- 
builders. Pentose also is important since it is the carbohydrate nucleus 
in the nucleo-proteid molecule of certain organs, the pancreas, thyroid, 
thymus, brain, spleen and liver. 

In the following paragraphs we shall quote the literature of this subject, 
warning the reader, however, that although the observations made by 
many may be accurate yet the conclusions of those who would interpret 
these observations as proving that the sugars in question were pentoses 
may not have been sufficiently confirmed. 

Pentosuria is said to occur under 3 conditions. First, an alimentary 
pentosuria will in normal persons follow the ingestion of considerable 
amounts of fresh fruit juice, or of beer. In these cases it is always an 
optically active pentose which is eliminated. Of all sugars the pentoses 
have the lowest assimilation limit. A dose of even 50 mgms. of pure xylose, 



184 CLINICAL DIAGNOSIS 

e.g., will produce a pentosuria. Second, some diabetic patients have 
pentosuria as well as glycosuria. It is only in very severe cases that the 
inability to burn sugars extends also to pentose (Kulz and Vogel found 
pentoses in the urines of 64 of 80 cases) . V. Jaksch found that diabetics 
excrete from 48.98 to 82.02% of the arabinoses, but a trace of the xylose 
and from 3 to 13% of rhamnose of the food, while non-diabetic patients 
excrete from 1 to 46.65% of arabinose, from 54.8 to 18.7% of xylose and 
from 63 to 55% of the rhamnose of the food. 

Third, there are on record a few cases of idiopathic pentosuria, a condi- 
tion quite different from diabetes. This was first discovered by Salkowski 
and Jastrowitz in 2 cases of suspected glycosuria. In 1902 only 5 or pos- 
sibly 6 70 such cases had been reported. Bendix later collected 12 cases 
and added 1 . It is interesting that several of these were chronic morphia 
habitues. In 1 of these the pentosuria continued after the habit was 
cured, but did not in another. 

Janeway 71 collected 22 cases of pentosuria and added 2 who interest- 
ingly enough were brothers. 

Pentosuria would seem to be a chronic symptomless condition discov- 
ered usually by accident. The amount of pentose on the urine is not large. 
In Salkowski' s case the amount eliminated would correspond to from 0.07 
to 0.15% and in Bendix's case, to from 0.4 to 0.6% of xylose. Bial 72 
found that in such cases the assimilation limit for both glucose and pentose 
is normal. The only explanation for the condition that he can offer is 
that there is an excess of pentose formed. 

Urines containing pentose will reduce copper, but not readily. The 
routine tests would suggest that a trace only of glucose or lactose is present. 
The reduction develops slowly after cooling, and suddenly throughout 
all the urine. Such urines do not ferment, are slightly dextrorotatory, and 
give with Nylander's solution a gray precipitate. 

Xylose, the most important clinically of the pentoses, is dextrorotatory, 
does not ferment with gas production, forms osazones with a melting point 
of 1 59 to 160 C, reduces copper and Nylander's solutions, gives an orange 
precipitate with Ruberner's test, and positive furfurol reaction. 

The arabinoses reduce copper and Nylander's solutions somewhat 
better than does xylose, and form osazones, the melting point of which 
varies from 157 to 15 8° C. One of the arabinoses is dextrorotatory but 
it is the inactive arabinose which is of particular interest, since this is the 
sugar found in the urine of the idiopathic cases. 

The Pkloroglucin Reaction. — To a few cubic centimeters of the urine are 
added an equal amount of HC1 (sp. gr. 1.19) and then from 25 to 30 mgms. 
of phloroglucin. The solution is then warmed until it becomes red in color. 

70 Brat's case, Munch, med. Wochenschr., 1903. 

71 Am. Jour. Med. Sc, Sept., 1906. 

72 Verh. d. xix. Kongr. f. inn. Med. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 185 

This solution if examined at once spectroscopically will show a band in the 
green if the pentoses are present. 

Salkowski recommends the following modification of this test: Five 
to six cubic centimeters of fuming HC1 are warmed and saturated with 
phloroglucin, leaving some undissolved. This solution is then equally 
divided in 2 tubes. To the 1 is added 0.5 c.c. of the suspected urine, and 
to the other 0.5 c.c. of normal urine. Both test-tubes are then placed in a 
beaker of boiling water. The tube containing pentose will soon be red in 
color, the change in color beginning above and extending downward. The 
tubes should be removed from the water-bath as soon as the red color 
begins to appear, and the red fluid examined spectroscopically. 

To exclude glycuronic acid, the pentosazone must be obtained (Sal- 
kowski). To from 200 to 500 c.c. of urine in a beaker are added 2.5 gms. 
per 100 c.c. of phenylhydrazin dissolved in an excess of acetic acid (or 
3.5 gms. HC1 phenylhydrazin with 1.5 parts of NaAc). This fluid is then 
warmed to the boiling point, the beaker allowed to stand in boiling water 
from 1 to 1% hours and then cooled. If pentose is present in any consider- 
able amount an abundant sediment of crystals will separate out. As soon 
as the crystallization is complete the precipitate should be recrystallized 
from a hot, very dilute alcoholic solution, and this repeated until the 
melting point is constant. 

The orcin-HCl test is to be preferred since it is somewhat more specific. 
To the urine, decolorized with animal charcoal, is added an equal volume 
of concentrated HC1, and then a small amount of orcin. This solution is 
then heated. If pentose or glycuronic acid (liberated from its paired com- 
pound by the acid and heat) is present, the color of the fluid turns to a 
reddish-blue (although the red may be very transitory and sometimes 
not appear at all) and finally become green. The solution is then cooled 
until it is only warm, and is then shaken out with amyl alcohol. A green 
fluid with a characteristic absorption spectrum is obtained. 

BiaVs modification of the Salkowski-Blumental test 73 is as follows: A 
reagent (HC1, 30%, 500 c.c; orcin, 1 gm. ; 10% Fe 2 Cl 6 , 25 drops) is used. 
Four or five cubic centimeters of this reagent are heated to boiling, then 
removed from the flame and the urine under examination added drop by 
drop, the total amount added not to exceed 1 c.c. If pentose is present a 
fine green color will soon appear. Bial claimed that this test excludes the 
glycuronic acid compounds, in fact is given by pentose alone. 

If hexoses also are present in the urine they, together with the pentoses, should be 
precipitated as osazones and the separation then made. The attempt should not be 
made to remove them by fermentation since the pentoses also will disappear during the 
process possibly as the result of bacterial action. 

Method of Kulz and Vogel. — From 1.6 to 3.2 liters of urine are used. To it are added 
200 gms. of phenylhydrazin plus 100 gms. of glacial acetic acid for each 100 gms. of 

73 Deutsch. med. Wochenschr., July 2, 1903. 



186 CLINICAL DIAGNOSIS 

glucose in the urine. The urine thus treated is then heated on a water-bath for an hour 
and a half, cooled and filtered. The filtrate is again heated on the bath for V/ 2 hours and 
filtered. These combined precipitates are well washed with cold water and then 
digested in water at 6o° C, which will dissolve the pentosazone. (Glucosazone can be 
dissolved only if the heat is raised to the boiling point.) For this digestion of the crystals 
I liter of water per ioo gms. of sugar is used, and the digestion continued 12 hours. 
This is repeated 15 times. The hot extracts are then filtered and allowed to cool, during 
which process the pentosazone will separate out. This precipitate is :repurified, using 
less water, until the melting point of the crystals is constant. 

The separation of the different pentoses is made with the polariscope. 
The alcoholic solution of zylosazone will show a strong constant levorota- 
tion, while arabinosazone immediately after formation is dextrorotatory 
and later becomes optically negative. 

One reason why cases of so-called pentosuria would seem to be rare 
may be that they are overlooked, or, unfortunately for the patients, usually 
diagnosed as glycosuria. But it is also true that pentosuria may prove 
to be a term covering a heterogeneous group of cases the urines of which 
give reactions suggestive of pentose, but, as certainly is the case, in some 
it is not pentose. 

Inosite. — Inosite is a carbohydrate-like body (really a hexahydroxylbenzol, 
C6H 6 (OH) 6 H 2 0) which is widely distributed in the vegetable kingdom. It is sometimes 
present in small amounts in the urine of patients with nephritis and diabetes and other 
conditions with polyuria. Naunyn mentions a case, probably of diabetes insipidus, who 
eliminated 18 to 20 gms. of inosite per day. Hoppe-Seyler believed that it may be found 
in all normal urines. 

The albumin should first be removed, the urine evaporated to % its volume and then 
precipitated with baryta water. The precipitate is washed, decomposed with H 2 S and 
filtered. The filtrate is allowed to stand in order that the unc acid may precipitate 
and be removed by filtration. The filtrate is then concentrated to a syrup and treated 
while boiling hot with 2 to 3 volumes of alcohol. A precipitate rapidly forms. This is 
cooled and ether is added after which the crystals of inosite will slowly appear. These 
are then purified by decolorization and recrystallization. 

Scherer's Test. — Inosite is evaporated to dryness with nitric acid on a platinum foil. 
To the residue are added a little ammonia and 1 drop of CaCh. It is then again evapor- 
ated to dryness and a fine rose-red residue obtained. This test is satisfactory only 
when the inosite is fairly pure. 

SeideVs Test. — This is the same as the above except that strontium acetate is used 
instead of CaCl 2 . A fine green colored solution with a violet precipitate develops. This 
test is positive even when but 0.3 mg. of inosite is present. 

Gallois Test. — The inosite solution is evaporated almost to dryness and the residue 
moistened with a little mercuric nitrate. On drying the solution a yellow residue is 
obtained, which at high heat becomes of a fine red in color, which color disappears on 
cooling and reappears on warming. 

Glycogen (Erythrodextrin) . — This has been found in the urine of diabetics after 
the glucose has disappeared or diminished, as a dextrin-like substance which browns on 
the addition of iodine. Urine containing it reduces copper after long boiling. To detect 
glycogen the urine is evaporated to a syrup, and KOH and absolute alcohol added -until 
a cloud due to the potassium salts appears. The fluid is then decanted, the precipitate 
is washed several times with absolute alcohol, dissolved in acetic acid, and reprecipitated 
with absolute alcohol. This precipitate is warmed with the alcohol and dried. A white 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 187 

tasteless powder is obtained, soluble in water, which reduces copper slowly and browns 
on the addition of iodine. 

Animal Gum (Landwehr). — Animal gum is said to be present in normal urine. It 
seems to be of the nature of a pentose, is slightly dextrorotatory, and does not ferment 
with gas production. With the copper test one obtains a precipitate which on boiling 
does not blacken. Alfthan found it much increased in the urine in diabetes mellitus 
(1.2 to 36.9 gms. per day; normally from 0.1 to 0.2 gm.). It is probably not 1, but a 
group of bodies precipitable by alcohol. 

Laiose is a sugar the nature of which is uncertain, found by Leo 74 in the urine 
of some, but not of all, diabetics. It is levorotatory, non-fermentable, with a salty 
taste and little reducing ability except after long boiling. It gives an oily compound 
with phenylhydrazin. 

Maltose was present in small and varying amounts in the urine of two patients. 

Isomaltose. — This sugar has been demonstrated in normal urine. Whether pre- 
formed or formed from glucose is uncertain. Certainly this transformation is very 
easy. The osazone precipitates in the form of very fine crystals which have a melt- 
ing point of 150 to 1 53 C. It either does not ferment or if it does it is very slowly, 
it reduces copper and bismuth and is dextrorotatory. It may be demonstrated as a 
benzoate compound. 

Melituria. — In the case of some malingerers it may be necessary to test the urine 
for cane sugar. In Brown's 75 case the urine had a high specific gravity, gave a positive 
but unsatisfactory Fehling's test, and but very few crystals with phenylhydrazin. It 
fermented very slowly. It was dextrorotatory. These tests would suggest that there 
may have been a trace of glucose in this urine, but it is more likely that some of the cane 
sugar had been inverted by the acid or by the bacteria of the urine. To demonstrate 
cane sugar the urine is concentrated, boiled with dilute HC1 for from 20 to 40 minutes 
and neutralized with sodium bicarbonate, after which the typical tests of glucose and of 
levulose may be obtained. 

Acetone. — There is some acetone in all normal urines, the maximum 
normal output being about 10 mg. in 24 hours. It is much increased in the 
following conditions: 

(1) Alimentary Disturbances. — The acetone of the urine is increased 
whenever the carbohydrates of the diet are much reduced. It is always 
increased by a diet rich in proteid and hence also during hunger periods. 
It is increased also by a diet rich in fat, although it required the ingestion 
of about 150 gms. of fat to influence the output. That an acetonuria may 
be of gastro-intestinal origin and is often associated with indicanuria is 
suggested by Wilson. 76 

(2) Febrile Acetonuria. — An acetonuria may be met with in any case 
of fever, no matter how slight. It has no clinical importance. 

(3) Diabetes. — In diabetes the output of acetone in the urine is the 
largest of all conditions. An intense acetonuria means an advanced, a 
long-standing case, and usually a severe case. While there are exceptions 
to this rule and while its presence need not necessarily mean an unfavorable 
prognosis, the curve of the acetone output is of some value in following an 

74 Virchow's Arch., 1887, vol. cvii, p. 108. 

75 Johns Hopkins Hosp. Bull., May, 1900, p. 101. 

76 The J. of Lab. and Clin. Med., May, 1920, v. 515. 



188 CLINICAL DIAGNOSIS 

acidosis. A severe case may eliminate more than 5 gms. of acetone daily. 
The acetonuria may increase greatly following slight fever; it is decreased 
by the administration of alkalies ; the output of cases on rigid diet may be 
reduced by adding carbohydrate to the food. Finally, it tends to increase 
as coma develops and toward death. Traces of acetone are eliminated also 
by the breath, 150 mgms. even being excreted in 1 hour through the lungs. 

Folin 77 has upset a few of the generally accepted ideas concerning ace- 
tone. He finds that its amount in the urine of even severe cases of diabetes 
is very small indeed; that the most of the substance estimated as acetone 
is diacetic acid; that some of the tests of acetone, e.g., Legal's, are really 
very delicate tests of diacetic acid; and that while the diabetics' breath 
and urine may contain acetone and may have a fruity odor, yet this odor 
is not due to acetone. 

(4) Patients with carcinoma in which inanition has not yet begun. 
(5) Cases of inanition and cachexia. (6) Psychoses and lesions of the 
central nervous system, especially those associated with starvation. (7) 
" Autointoxication." (8) Digestive disturbances, especially gastric ulcer. 
(9) Chloroform narcosis, in which case it is due to the increased proteid 
catabolism. (10) Pregnancy with a dead fetus. (11) Certain poisons: 
e.g., phloridzin. (12) Extirpation of the pancreas. 

There is some doubt that there is any preformed acetone in the body. 
All in the urine would seem to be derived from diacetic acid. 

Acetone, CH3COCH3, is a thin, colorless fluid with a specific gravity 
of 0.814 (at o° C.)i a boiling point of 56.5 C. and a quite characteristic odor. 

Tests. — As a general rule only the distillate of the urine should be 
tested for acetone. From 250 to 1000 c.c. of fresh urine are used and a 
little acid, preferably phosphoric, added to prevent foaming. V. Jaksch 
advises that it be distilled with steam, in which case no acid is necessary. 
A good cooler should be used if the acetone is to be determined quantita- 
tively, although for qualitative work this is not necessary. Most of the 
acetone will pass over in the first 10 to 30 c.c. of the distillate. Since 
diacetic acid is easily split up to acetone the urine should first be made 
alkaline and carefully shaken out with alcohol-free ether if it is desired 
(as it seldom is) to exclude this body. This ether extract may then be 
shaken out with water and the latter tested for diacetic acid. 

Legal's Test. — As a preliminary test, Legal's may be used and yet this 
is satisfactory only when large amounts of acetone are present. A negative 
result has little value. 

To the urine or its distillate are added a few drops .of fresh concentrated 
solution of sodium nitroprusside, and then KOH or NaOH until the 
reaction is very alkaline. A ruby-red color appears which changes rapidly 
to yellow. The test thus far is the same as that given by creatinin. Gla- 
cial acetic acid therefore is added in excess to the still red fluid. If acetone 

77 Jour, of Biol. Chem., May, 1907; Jour, of A. M. A., May 2, 1908. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 189 

is present the red will change to a purple-red and later to a violet color. 
Creatinin would give a yellow color which would change to green and finally 
to blue. Paracresol gives a reddish-yellow solution; acetic acid a clear 
rose color. To exclude aldehyde Le Noble and Lee used NH 4 OH in place 
of KOH. 

Gunning's test is very satisfactory and accurate. To the distillate is 
added tincture of iodine or Lugol's solution (KI, 1.8; I, 1.2; H 2 0, q. s. 
ad 30), and then ammonia until a deep black precipitate forms. This 
later gradually disappears, leaving a yellow sediment of hexagonal or star- 
shaped iodoform crystals which may be recognized by their color, odor and 
shape (see Fig. 33). The sediment is seldom amorphous. In case but a 
trace of acetone was present it may be necessary to wait 24 hours before 
the sediment appear. If necessary the iodoform may be recrystallized 
from ether. Gunning's test is less delicate than Lieben's, but is given by 
no other body than acetone, of which it will show 0.01 mgm. per 1 c.c. 

Denige's test 78 is preferred by some : To about 
Y 2 inch of the distillate of the urine in a test-tube 
is added an equal amount of a solution of the 
subsulphate of mercury (mercuric oxide, 50; sul- 
phuric acid, 200; water to 1000). This mixture 
is allowed to simmer about 5 minutes. When 
it cools there is deposited a white crystalline 
precipitate, which is distinctive in appearance, 
and is not soluble in dilute HC1. 

A test now popular is as follows: 79 To 10 c.c. 
of urine in a test-tube one adds about 1 gm. of 
ammonium sulphate, 2 to 3 drops of a freshly Fo^ed^f^lh^stinSf^ 
prepared 5% solution of sodium nitroprusside, the urine of a case of dlabetes " 
and 2 c.c. of concentrated ammonium hydroxide which may be stratified 
or poured on the mixture. The presence of acetone is indicated by the 
slow development of a permanganate color. The test is positive if there 
is 1 part of acetone in 20,000 parts of urine. 

Quantitative Determination of Acetone Plus Diacetic Acid. — 
The Huppert-Messinger Method. By this method one determines the 
sum of the acetone and of the diacetic acid which is transformed to acetone 
by the process of distillation. 

The solutions necessary are: 

1. Acetic acid, 50%. 

2. 0.1 N iodine solution. 

3. 0.1 N sodium thiosulphate solution. 

4. A thin starch solution (1 gm. of starch dissolved in 500 c.c. of boil- 
ing water) . 

78 See Taylor, Jour, of A. M. A., March 17, 1906, vol. 46, p. 790. 

79 Wilson, Jour, of Clin, and Lab. Med., 1920, v. 515. 



JTT- 7 


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190 CLINICAL DIAGNOSIS 

To make up these solutions 24.8 gms. of crystallized sodium thiosulphate 
(Na 2 S 2 035H 2 0) are carefully weighed, dissolved in distilled water, and the 
solution made up accurately to 1 liter. Next 2 5 gms. of potassium iodide 
are dissolved in a little water, 12.7 gms. of iodine added, and the solution 
made up to about 900 or 950 c.c. To standardize this solution, 20 c.c. 
of the thiosulphate solution are carefully measured into a small flask, a 
few drops of the starch solution added, and then the iodine' solution run 
in from a buret with glass stop-cock until the blue color just appears. 
This titration is repeated several times until the amount necessary for the 
end reaction is accurately determined. Then to the iodine solution is added 
the necessary amount of water so that 20 c.c. of this solution will exactly 
equal 20 c.c. of the thiosulphate solution and the resulting solution con- 
firmed by another titration. 

Both of these fluids are to be kept in dark glass bottles with ground-glass 
stoppers. The iodine solution must be restandardized frequently. One 
cubic centimeter of the thiosulphate solution equals 0.0127 gm. of iodine. 
The formula of the reaction is 2l + 2Na 2 S 2 3 = 2NaI+Na 2 S40 6 . The first 
trace of free iodine in excess will form the blue starch-iodine compound. 

The urine if alkaline is first made just acid with acetic acid. To 500 c.c. 
of acid urine (if rich in acetone use 100 c.c. or even less) is added exactly 
2 c.c. per 100 c.c. of the urine of 50% acetic acid. The urine is then distilled 
into a flask surrounded by ice and tightly closed by a stopper with two 
perforations. Through 1 of these passes the tube from the distilling flask, 
which tube reaches to the bottom of the flask and dips below the surface 
of water previously placed there; through the other perforation passes a 
shorter tube connected with a bulb or Peligot U-tube filled with water, 
which acts as a safety bulb to prevent the loss of any of the very volatile 
acetone. Then the urine is distilled until about % of its volume has passed 
over into the receiving flask. The distilling flask must be disconnected 
before the heat is removed, else the distillate will " strike back." The tube 
of the cooler is now washed thoroughly with distilled water over into the 
receiving flask so as to conserve the last trace of distillate. Lastly and for 
the same reason the water in the safety bulb or U-tube is emptied into, 
and its tube washed into, this flask. 

Some calcium carbonate is then added to the distillate and the flask 
well shaken. This will remove any nitrous and formic acids which may have 
distilled over. 

The distillation is now repeated as before. To this second distillate 
(to which again is added the water in the safety tube and the wash-water 
from both tubes) is added 1 c.c. of dilute sulphuric acid (1 : 8 of water), 
and the distillation again repeated, using the same precautions as before. 

This final distillate is poured into a flask or measuring cylinder with 
ground-glass stopper and so large that the distillate and reagents next to 
be added will not fill it more than % full. (Or, this flask or cylinder may be 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 191 

used to receive the distillate during this distillation. It should of course 
be protected as in the first distillation.) A large excess of carefully meas- 
ured o.i N iodine solution is added, these fluids well shaken, and then an 
excess of strong nitrate-free NaOH or KOH added drop by drop. The 
flask is closed, shaken for % of a minute and then allowed to stand for 
5 minutes. 

The stopper is then removed (and any fluid clinging to it washed back 
into the flask), the contents of the flask are then made acid with concen- 
trated hydrochloric acid and the excess of iodine determined by titration 
with the o.iN thiosulphate solution which should be added from a buret 
until the mixture is slightly yellow. Then a few cubic centimeters of the 
starch solution are added and the titration continued until the blue color 
has just disappeared. If one accidentally carries the titration too far he 
may add more of the iodine solution, carefully measured, and then continue 
the titration until the end reaction is reached. One cubic centimeter of 
the iodine solution indicates 0.967 mg. of acetone. 

The results of this method are from 4 to 8% too low. The final distillate 
must contain no phenol, ammonia, nitrous or formic acids, for all of these 
but nitrous acid will cause the loss of some of the iodine, while nitrous acid 
will set iodine free. It is to prevent the error from ammonia that 2 c.c. of 
acetic acid per 100 c.c. of urine were added to the just-acid urine. If a 
mineral acid or as much as 5 c.c. of acetic per 100 c.c. of urine were used 
none of the ammonia would reach the distillate, but some phenol would. 
For this reason only 2 c.c. of acetic per 100 c.c. of urine are added and the 
trace of ammonia which does distil over is later removed by the third 
distillation after the addition of sulphuric acid. The addition of calcium 
carbonate to the first distillate will remove the nitrous and formic acids. 

It is wise not to shake the final distillate when one adds to it the alkali 
and the iodine solution and to notice whether a black color appears at the 
line of separation of these 2 fluids. If it does, ammonia is present and the 
specimen should be thrown away. If not, the fluid is shaken, and one pro- 
ceeds as above. 

A much easier and fairly satisfactory method of determining the sum 
of acetone and diacetic acid in term's of acetone is the following : 

From 50 to 250 c.c. of urine, according to the amount of acetone, to 
which is added a little acid to prevent foaming, is measured into a distilling 
flask. To the end of the outlet tube, which should pass through a very 
efficient cooler, is attached a rubber tube, the end of which dips beneath 
the surface of water previously placed in the receiving flask. The distilla- 
tion is continued until most of the water has passed over. 

The distillate is poured into a graduated cylinder with a ground-glass 
stopper, an excess (15 to 20 c.c.) of NaOH added, and then 20 c.c. of Lugol's 
solution, which may conveniently be made 3 times the ordinary strength. 
A heavy black precipitate forms which soon clears, leaving a yellow sedi- 



192 CLINICAL DIAGNOSIS 

ment of iodoform crystals. After standing for 10 to 15 minutes or more, 
about 40 to 50 c.c. of ether are added and the fluid well shaken until the 
ether contains practically all of the iodoform. After a reading is made of 
the volume of ether, 10 c.c. of it are measured with a graduated pipet 
into a weighed glass dish and evaporated in the air. This is then dried 
to constant weight over sulphuric acid. The weight of the iodoform multi- 
plied by 0.147 equals the weight of acetone represented in the 10 c.c. of 
ether extract used. From this the amount in the entire volume of ether 
extract may be reckoned. 

Faun's 80 method allows an estimation of the acetone alone. The acetone 
is separated from the urine by the same apparatus devised by Folin for 
ammonia estimations (see page 124). The urine, 25 c.c, is measured into 
the aerometer cylinder, and from 0.2 to 0.3 gm. of oxalic acid or a few drops 
of 10% phosphoric acid and 8 to 10 gms. of sodium chloride and a little 
petroleum added. In the receiving bottle has been previously placed 
water to which is added 40% potassium hydroxide solution (10 c.c. per 
150 c.c. of the water) and an excess of the standardized iodine solution. 
The apparatus is then connected with a Chapman air pump and a fairly 
strong (yet not as strong as for an ammonia determination) air current 
drawn through for from 20 to 25 minutes. Every trace of acetone will be 
removed from the urine and converted in the receiving bottle to iodoform. 
The contents of the receiving bottle are acidified with concentrated hydro- 
chloric acid (10 c.c. for each 10 c.c. of alkali used) and the excess of iodine 
titrated with standard thiosulphate and iodine solutions as in the Messinger 
method. The observer must be thoroughly acquainted with his apparatus 
and the strength of his air current by repeated experiments, using solutions 
to which known amounts of pure acetone have been added. 

Diacetic Acid — Acetoacetic Acid, CH3COCH2COOH. — Diacetic acid 
is from the clinical viewpoint the most important of the acetone bodies, 
since it not only is the easiest to test for, but also is a very important index 
of acidosis. The present idea is that all diacetic acid in the urine is derived 
from /3-oxybutyric acid, and all the acetone in turn from diacetic acid. 
The urine of a normal person should contain but a trace, if any, of diacetic 
acid and probably none if on a mixed diet. It appears quite early in the 
urine of a person starving, or on a diet poor in carbohydrates, and will 
promptly disappear if even a little carbohydrate be added to the diet. 
But individual differences in the output of this acid are so great that 
some other factors must be more important than the mere lack of carbo- 
hydrate in the diet. The statement is often made that one finds diacetic 
acid in those urines only which contain acetone in large amounts and not 
always then, yet the work of Folin would seem to prove that acetone is 
never present in such urines in more than traces, and that the tests used 
for acetone show in fact diacetic acid. 

80 The Jour, of Biol. Chem., May, 1907. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 193 

Diacetic acid is found in the urine in increased amounts in the same 
conditions mentioned in page 187 for acetone. That is, in diabetes mellitus, 
in conditions of undernutrition or with defective absorption from the bowel, 
and hence in all cachexia-producing diseases; and in fevers, even in mild 
cases, especially the acute exanthemata of children during the eruptive 
stage and still more especially in streptococcus infections. Its amount 
would seem to depend much on the nature of the infection. Its presence 
is not limited to the febrile periods and it has no prognostic importance. 
Next to diabetes the most important group of cases with diacetic acid in 
the urine is that of gastro-intestinal disturbances. In these cases its pres- 
ence is by no means explained by the lack of carbohydrate, but would seem 
to depend on abnormal fat catabolism. It is found in very mild cases; it 
does not disappear when sugar is added to the diet as promptly as it does 
from the urine of a normal person on a pure proteid diet. It is said to occur 
in especially large amounts in the urine of drunkards with gastro-intestinal 
disturbances. Rolleston and Tebbs 81 found it present in abundance in 
33 of 38 cases of gastric ulcer treated either by starvation or by rectal 
feeding. The tests for it became positive in from 2 to 12 days, usually 
1 or 2 days, after treatment began and disappeared in from 1 to 14 days, 
usually 5, after the return to mouth feeding. Women would seem to 
excrete especially large amounts of this acid; age and the chronicity of the 
disease would seem of no moment in its production. In some of these cases 
as much ammonia (the best index of acids) is present in the urine as in 
diabetes (Golla). It may be found in the urine of normal men who for a 
few days have been on a pure proteid diet, and in mental cases who are 
losing weight and suffering from inanition. 82 

Gerhardt's Test. — The best test for diacetic acid is that proposed 
by Gerhardt. To from 10 to 50 c.c. of fresh urine are added a few drops 
of a Fe 2 Cl 6 solution, which must not be too acid. This is added as long as 
a precipitate forms, and then the urine is filtered. To the filtrate is added 
still more Fe 2 Cl 6 . If diacetic acid is present the urine takes on a Bordeaux- 
red color, cherry-red by transmitted, purple-red by reflected, light. This 
test indicates from 0.4 to 0.5 p.M. of diacetic acid. Cyanates, NaAc, sali- 
cylic acid and its allied bodies, salol, aspirin, diuretin, and certain other 
medicines also will give a somewhat similar color, but with this difference 
that of all these substances diacetic acid is the only one which is destroyed 
by heat, hence a positive test should always be controlled by another made 
after the urine, first made weakly acid, has been boiled and then cooled. 
In this control test the red color of the urine should be distinctly paler 
than in the unboiled, since some at least of the diacetic acid will have been 
broken down. A modification of this test is to shake out the urine acidified 
with H2SO4 with ether and this with water, and to add the Fe 2 Cl 6 solution 

81 Brit. Med. Jour., 1904, vol. ii, p. 114. 

82 See also Futcher, Med. News, Oct. 8, 1904. 

13 



194 CLINICAL DIAGNOSIS 

to the water extract. A violet-red color appears in the water layer if this 
acid is present. This color pales on standing in from 24 to 48 hours, a 
necessary part of the reaction to exclude other bodies. 

Arnold 's Test. — Two solutions are kept in stock. One is a 1 % aqueous 
solution of para-amido-acetophenon with 2 c.c. of strong hydrochloric acid 
in each 100 c.c. of the mixture; the other is a 1% solution of potassium or 
sodium nitrite. Two parts of the first and one of the second solution are 
mixed together in a test-tube, an equal bulk of urine added and finally a 
drop of strong ammonium hydrate. In normal urines a brown color usually 
appears which on the addition of several drops of strong hydrochloric acid 
changes to yellow, while in a urine containing diacetic acid a purple color 
develops. Normal urines may develop a red color similar to that given 
by urines containing very slight traces of diacetic acid, but on shaking 
the urine containing the diacetic acid the foam will show a violet color. 

/3-oxybutyric Acid, /5-hydroxybutyric Acid, CH 3 CHOHCH 2 COOH.— 
Butyric acid is considered the mother substance of diacetic acid and hence 
of acetone. One might expect therefore to find it when they are present, 
especially if they are present in large amounts. Yet the chances are against 
finding it since it breaks up so readily to form diacetic acid. Gerhardt 
and Schlesinger 83 showed that it will appear in the urine of a normal man 
who has been for some days on a proteid diet (about 9 gms. of this acid 
were eliminated in 24 hours). This is the acid to which is attributed the 
acid intoxication (or alkali starvation) which is said to explain diabetic 
coma. Often about 50 gms. a day are excreted, and in one of Naunyn's 
cases 100 gms. a day for a long time. Larger amounts are excreted during 
coma: 188 gms. in 24 hours (Magnus-Levy) 5225 gms. (Kulz) and in Joslin's 
case, 437 gms. in 3 days, i.e., 3 gms. per 1 kilo per day. 

Joslin believes that it is the alkaline treatment itself which explains 
these huge figures and cautions against the administration of any alkalies 
at all. Whether oxybutric acid has a specific toxicity or is toxic because 
it is an acid, is in doubt. The Strassburg school holds the latter view. On 
the other hand Wilbur 84 found that the injection into animals of this 
neutralized acid gave results similar to those of the free acid. He empha- 
sized the point that the alkaline treatment of coma has not been as satis- 
factory as one would on theoretical grounds expect. 

This acid is levorotatory, [a]D= — 24.12 . Its presence, therefore, 
may be suspected when the percentage of sugar measured with the polari- 
scope is less than that indicated by titration. The presence of other 
levorotatory bodies, however, should always be considered: levulose, 
paired glycuronic acid compounds, albumin, etc. 

Detection. — |8-oxybutyric acid is probably present if the fermented 
urine of a diabetic shows a definite levorotation. It is quite certainly 

83 Arch. f. exp. Path. u. Pharm., 1898. 

84 Jour. Am. Med. Assoc, 1904, No. 17. 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 195 

present if the ether extract of urine which has been fermented, then acidified 
with phosphoric acid and then extracted with ether, is levorotatory. 

A very reliable test for this acid is the following : The urine is fermented 
till all glucose is gone. It is then evaporated to a syrup, an equal amount 
of concentrated H2SO4 added and then distilled. Crotonic acid is now 
present in the distillate which when cooled will separate in beautiful 
crystals with a melting point of 72 C. If these crystals do not readily 
form, the distillate may be shaken out with ether, the ether evaporated, 
the residue washed with water and then allowed to crystallize. 

Quantitative Determination — Black's Method. — One measures 100 
c.c. of urine with a pipet into an evaporating dish, makes it distinctly 
alkaline with sodium bicarbonate and then evaporates it to a thick syrupy 
liquid (4 or 5 c.c), using the gentle heat of a water-bath or the low heat 
of an electric stove. This residue is cooled, made distinctly acid with strong 
hydrochloric acid and is then mixed gradually with plaster-of-Paris until 
it forms a porous, mealy mass. This porous meal is transferred to an 
extraction apparatus (that of Soxhlet or one of its modifications) and 
extracted for 3 hours with about 60 c.c. of ether. The ether extract is 
then transferred to an evaporating dish and the ether allowed to evaporate 
spontaneously. The residue is mixed with 5 c.c. of water, 0.4 gms. of 
bone-black added to decolorize it and it is then filtered and washed until 
perfectly clear. The filtrate is next made up to a known volume, usually 
25 c.c, and the amount of /3-hydroxybutyric acid which it contains deter- 
mined with a polariscope, using the following formula : 

>, . . . - . Angle observed 

Grams of /3-oxybutvnc acid in 1 c.c. = 

24.12X200 

24.12 = specific rotation of this acid 

200 = length of polariscope tube. 

Shaffer's Method for Determining (3-hydroxybutyric Acid, Acetone and 
Diacetic Acid Combined. — One measures from 25 to 250 c.c. of the urine 
to be examined (estimating as nearly as possible that volume which would 
contain from 25 to 50 mgms. of acetone — from 25 to 50 c.c. of a urine 
which gives a strong ferric chloride reaction will suffice) and adds to it a 
definite excess of basic lead acetate and 10 c.c. of ammonium hydroxide. 
The mixture is made up to the 500 c.c. mark, shaken thoroughly and filtered 
through a dry filter paper. To 200 c.c. of the filtrate, measured into a dis- 
tilling apparatus, are added from 300 to 400 c.c. of water, 15 c.c. of con- 
centrated sulphuric acid and a little talcum to prevent knocking, and 
the distillation continued until from 200 to 250 c.c. of distillate have 
collected. This distillate contains, in addition to the preformed acetone, 
that which results from the breaking down of diacetic acid and certain 
volatile acids. It is therefore made slightly alkaline and distilled a 
second time. This distillate will contain all of the acetone (that originally 



196 . CLINICAL DIAGNOSIS 

preformed and that from the diacetic acid) and this may be estimated by 
the iodine method. 

The residue containing the sulphuric acid contains all of the 0-hydroxy- 
butyric acid, which now must be converted by oxidation into acetone. The 
residue is therefore diluted, 0.5 gm. of potassium dichromate added and 
then distilled from a flask provided with a dropping funnel through which 
water is added gradually to make up for the amount distilled over. Instead 
of water one may use a 0.5% solution of potassium dichromate. To about 
500 c.c. of the distillate are added 20 c.c. of 3% hydrogen dioxide, sufficient 
potassium hydroxide to make the solution alkaline and the distillate is 
redistilled until about 300 c.c. have passed over. The amount of acetone 
in this distillate is estimated by the iodine method. One milligram of ace- 
tone is equivalent to 1.794 mgms. of /3-hydroxybutyric acid. 

Van Slyke-Fitz Methods for the Determination of (3-hydroxybtityric Acid, 
Diacetic Acid and Acetone in Urine and Blood (quoted from Joslin) — Solu- 
tions Required. — A 20% copper sulphate solution prepared by dissolving 
200 gms. of Q1SO45H2O in water and making the solution up to 
1 liter. 

A 10% mercuric sulphate solution prepared by dissolving 73 gms. of 
chemically pure red mercuric oxide in 1 liter of 4N H 2 S0 4 . The solution 
of the oxide is assisted by warming it on a steam-bath. 

Fifty volume per cent, sulphuric acid prepared hj diluting 500 c.c. of 
H2SO4 (sp. gr. 1.835) to 1 liter with water. This should be 17N H 2 S0 4 . 

Colloidal iron; Merck's " dialyzed iron " solution, containing 5% 
of Fe 2 3 . 

Ten per cent, calcium hydrate suspension containing 100 gms. of 
Merck's fine light " reagent " Ca(OH) 2 mixed with 1 liter of water. 

Five per cent, potassium dichromate solution prepared by dissolving 
50 gms. of K 2 Cr 2 07 in 1 liter of water. 

To remove glucose and other substances which would interfere, 25 c.c. 
of the urine to be examined are measured into a 250 c.c. measuring flask, 
100 c.c. of water and 50 c.c. of the copper sulphate solution added and the 
solutions well mixed. To this are then added 50 c.c. of 10% calcium 
hydrate. The solution is well shaken and tested with litmus. If it is not 
alkaline in reaction more calcium hydrate is added. The solution is then 
diluted to the mark and let stand for at least y 2 hour to allow the glucose 
to precipitate. It is then filtered through a dry folded filter. This proced- 
ure will remove all the glucose of an 8% or weaker solution, therefore urine 
containing more should be so diluted that the percentage of glucose may 
be less than 8%. To be sure that the nitrate is free from glucose a little is 
boiled in a test-tube. A yellow precipitate (of Cu 2 0) will indicate glucose; 
a white precipitate (of CaCOs) has no significance. 

To remove all proteins from the blood 10 c.c. of whole blood are meas- 
ured into a 250 c.c. volumetric flask half full of water, 50 c.c. of colloidal 



THE URINE: CARBOHYDRATES AND ALLIED BODIES 197 

iron are added, this mixed, then i c.c. of the saturated sodium sulphate 
solution added. The flask is then filled to the mark, shaken and the con- 
tents filtered through a dry folded filter. 

If plasma is to be examined the procedure is the same except that only 
8 c.c. are measured in a 200 c.c. flask and 15 c.c. of colloidal iron and 1 c.c. 
of saturated sodium sulphate are added. 

In the case of either whole blood or serum 1 2 5 c.c. of the filtrate, equiva- 
lent to 5 c.c. of the original sample, are taken for analysis. 

Simultaneous determination of total acetone bodies (acetone, diacetic acid 
and jS-hydroxybutyric acid) of the urine or blood in one operation. Into 
a 500 c.c. Erlenmeyer flask are measured 25 c.c. of the urine filtrate plus 
100 c.c. of water; or 125 c.c. of the blood filtrate plus 10 c.c. of 50% sulphuric 
acid and 35 c.c. of the 10% mercuric sulphate solution. The flask is then 
connected with a reflux condenser having a straight condensing tube of 
8 or 10 mm. diameter and heated to boiling. After the boiling has begun, 
5 c.c. of the 5% dichr ornate solution are added through the condenser tube 
and the boiling continued gently for i}4 hours. The precipitate which 
forms consists of the mercury sulphate compound of acetone, both that 
preformed and that formed by the oxidation of the hydroxybutyric acid. 
The precipitate is collected into a Gooch, or a medium density " alundum," 
crucible, washed with 200 c.c. of cold water followed by a little 95% alcohol, 
dried for an hour at no° and weighed. Several precipitates may be col- 
lected, one above the other, without cleaning the crucible. 

Acetone and Diacetic Acid. — These substances without the hydroxy- 
butyric acid are determined exactly as are the total acetone bodies, except 
that (1) no dichromate is added to oxidize the hydroxybutyric acid and 
(2) the boiling must continue for not less than 3 5 nor more than 45 minutes. 
Boiling for more than 45 minutes would split off a little acetone from 
hydroxybutyric acid even though no dichromate had been added. 

(3-hydroxvbtityric Acid in Urine. — The /3-hydroxybutyric acid alone is 
determined exactly as total acetone bodies except that the preformed 
acetone and that formed from the diacetic acid are first boiled off. To do 
this the 25 c.c. of urine filtrate plus 125 c.c. of water are treated with 2 c.c. 
of 50% sulphuric acid and boiled in the open flask for 10 minutes. The 
volume of solution left in the flask is measured in a cylinder, then returned 
to the flask and the cylinder washed with enough water to replace part of 
that boiled off and to bring the volume of the solution to 127 c.c. Then 
8 c.c. of the 50% acid and 35 c.c. of the 10% mercuric sulphate solution 
are added. The flask is connected under the condenser and the determina- 
tion is continued as above. 

^-hydroxybutyric Acid in Blood. — -The following procedure enables one 
to determine separately in a single sample of blood both the acetone plus 
the diacetic acid and the hydroxybutyric acid. The acetone and diacetic 
acid are precipitated as above described and the filtrate poured as com- 



198 CLINICAL DIAGNOSIS 

pletely as possible through the Gooch or alundum crucible into a dry 
receiving flask. Of this nitrate 160 c.c. are measured into another Erlen- 
meyer flask and 10 c.c. of water are added. The mixture is heated to boiling 
under a reflux condenser, 5 c.c. of dichromate solution are added and the 
determination continued as described for " total acetone bodies." 

In case only the /3-acid is to be estimated, or if enough blood is taken 
for 2 determinations, the slightly easier procedure used for -urine may be 
followed also with blood. 

Factors for calculating the acetone bodies in urine, when 2 5 c.c. of the 
filtrate-, equivalent to 2.5 c.c. of urine, and in blood when 135 c.c. of filtrate, 
equivalent to 5 c.c. of blood, are used for determination. One milligram 
of /3-hydroxybutyric acid yields 8.7mgms. of precipitate. One milligram 
of acetone yields 19.7 mgms. of precipitate. 

The amount of precipitate obtained from /3-hydroxybutyric acid there- 
fore corresponds to 79% of the acetone that would be obtained if each 
molecule of hydroxybutyric yielded a molecule of acetone. The oxidation 
is complete in 1 )4 hours and the conditions are so constant that duplicates 
usually check within 1%. 

DIABETES MELLITUS 

Diabetes mellitus is a disease the most important characteristic of 
which is a reduction of the ability of the tissue cells to use the glucose 
molecule. So far as we know all cases of long-standing (i.e., more than 
1 week) glycosuria are cases of diabetes mellitus, but the urine of a patient 
with diabetes mellitus may be sugar-free even for months, provided the 
carbohydrates in the diet do not exceed the limit of ability of the body to 
warehouse glucose. Indeed a case of diabetes mellitus may theoretically 
during his life never be a case of glycosuria except during the periods 
when his sugar tolerance is being tested. The kidneys and therefore the 
urine are only secondarily involved in this disease. The lessened ability 
of the tissue-cells to burn glucose would seem to depend either on a limita- 
tion of the supply of some substance in the blood which like a key is neces- 
sary to unlock the glucose molecules (although their ability to burn any 
other molecule may be quite unimpaired) or of a substance which when in 
combination with glucose renders this suitable for use by the tissue cells. 
This missing substance has been called the " internal secretion," the " pan- 
creatic amboceptor," etc. The student should bear in mind that in a well 
marked case of diabetes mellitus we are dealing with at least 6 different 
disease complexes, and that these while related are nevertheless in many 
particulars independent: first, the systemic disease, which in diabetes 
happens to affect the pancreas especially, e.g., cancer, lues, etc., diseases 
which apart from their relation to the pancreas have their own pathology 
and prognosis; second, this disease of the pancreas itself; third, the glyco- 
suria; fourth, the production of abnormal bodies, many of them acid in 



THE URINE: DIABETES MELLITUS 199 

nature, evidently products of the incomplete combustion of fats and 
proteids; fifth, conditions arising from the alkali starvation produced by 
the presence and elimination of these acid bodies; and sixth, various com- 
plications and sequelae, as gangrene, degenerations in the central nervous 
system and peripheral nerves, pyogenic infections favored by the lowered 
resistance of the tissues due to the diabetic condition, etc. Each one of 
this group of conditions gives rise to problems in diagnosis peculiar to itself. 

Cases of diabetes mellitus may be classified as severe, if they have a 
tolerance of from o to 10 gms. of carbohydrate; moderate, if they have a 
tolerance of from 10 to 50 gms. and mild if the tolerance is over 50 gms. 
Since practically every case may be made sugar-free by proper fasting and 
diet, the older definition of severe cases as those who cannot be made sugar- 
free on a carbohydrate-free diet can no longer be used. How severe a case 
may be can often be determined only after long observation and will 
depend less on the history of his glycosuria than on the prognosis of his 
underlying diseases. 

This is well illustrated by the case of Geyelin and Dubois 85 who was 
admitted with symptoms of most severe diabetes mellitus of 6 weeks 
duration and bordering on diabetic coma and discharged 3 months later, 
a mild case. 

The urine in diabetes mellitus is, as a rule, but not necessarily, increased 
in amount. This increase is often not marked unless the urine contains 
over from 2 to 3% of sugar, beyond which point the volume is roughly 
proportional to the amount of glucose present. Severe cases, that is, those 
excreting 5% or more of sugar, may void from 4 to 5 liters, even 10 liters, 
while there is 1 case on record who voided 28 liters of urine in 24 hours. 
Joslin's record case was a boy 10 years old who weighed 18.6 kilos and who 
in 16 hours voided 7200 c.c. of urine or 39% of his weight. On the other 
hand some patients void small volumes of urine with a high percentage of 
sugar. This is considered to indicate a good prognosis although a polyuria 
does not indicate a bad one. Naunyn reported 2 such cases, 1 with 1400 c.c. 
of urine containing 9% of sugar and with a specific gravity of 1.040; a 
second with 2800 c.c, 10.5% of sugar and specific gravity 1.047. Joslin 
mentions a case with 1035 c.c. of urine in 24 hours and 5.8% of sugar. 
In other cases the reverse is true and there is a polyuria with a low percent- 
age of glucose, but this is rare except in those cases in which an increasing 
acidosis accompanies a decreasing glycosuria and especially in cases which 
follow injury to the skull. In one such case the specific gravity was 1.003 
and sugar 1% (Naunyn's case). A similar condition may be met with in 
a diabetic who is developing chronic interstitial nephritis and in some cases 
of diabetes with marked asthenia. 

It is in diabetes mellitus that the specific gravity of the urine reaches 
record figures. As a rule it ranges between 1.030 and 1.040. In one of 

85 Jour. A. M. A., 1916, vol. 66, p. 1532. 



200 CLINICAL DIAGNOSIS 

Naunyn's cases it was 1.060, and he mentions one reported in which it was 
1.074. A polyuria is usually present if the urine in a case of this form of 
diabetes has a specific gravity of 1.030. This is the one condition in which 
there is both a high specific gravity and an increased amount of urine, 
but the specific gravity bears little relation to the latter. 

The chief sugar present is glucose, yet levulose, pentose and other 
carbohydrates may also be present; in rare cases levulose alone is found. 
There is an increase also of the unfermentable carbohydrates [a minimum 
of 20 gms. instead of 1.6 gms. (normal maximum, 5 gms.) per day]. 86 

The urine has a suggestive pale greenish-yellow color. It will ferment 
spontaneously, with the evolution of C0 2 and the deposit of a sediment. 
This fermentation may take place in the bladder and consume the sugar 
so that a sugar-free urine may be excreted. Again, the sugar may disappear 
by a fermentation without gas production. 

In testing the urine qualitatively for glucose it is sometimes important 
to choose the right specimen. If sugar is present in all voidings this is 
not important, but mild cases may eliminate little sugar and that only 
during a few hours of each day. Did we examine this one voiding there 
would be no doubt as to the presence of sugar, but if this voiding is mixed 
with all the other voidings of that day the solution of glucose may be so 
dilute that it will not give a positive test. For that reason we examine 
that specimen of urine voided from 4 to 6 hours after a noon meal which 
contains some carbohydrate. The maximum of sugar excretion comes 
late in the forenoon, even though the ingestion of carbohydrates extends 
equally over the whole day. There is another maximum, a somewhat less 
one, in the late afternoon (about 6 o'clock). The minimal excretion is 
early in the morning. In some severe cases the variations in sugar output 
are but little marked and in still more severe cases much more may be 
excreted during the night than during the day (note the resemblance to 
the excretion of water and solids in nephritis). In mild cases on a mixed 
diet the urine may be sugar-free during the night and reach even 3% during 
the day. In some cases sugar-free periods lasting for months will alternate 
with periods of glycosuria. It is thus evident that mild cases may easily 
be overlooked if but one specimen of urine is examined. The output of 
sugar is greater in hot than in cold weather. This is also true of the carbo- 
hydrate of normal urines. Cases of diabetes mellitus have been classified 
according to their percentage of sugar excretion (their " intensity ") and 
also according to the total amount of sugar excreted in a day (or their 
" size ") . Cases can be compared in this way only if they are on a constant 
diet and such a classification has but little value. The amount of sugar 
eliminated per day by severe cases on a liberal diet is often 800 gms. and 
in one case 1500 gms. in 24 hours. These same patients on a more limited 
diet seldom excrete over 200 gms. Starving patients are usually sugar-free. 
86 Edsall, Am. Jour. Med. Sci., 1901. 



THE URINE: DIABETES MELLITUS 201 

As regards the relation between water and glucose excretion, it may be 
said that diabetics responded to an increased intake of fluid more slowly 
than do normal people. In glucose the water excretion depends in large 
measure upon the amount of glucose to be eliminated although this does 
not explain the day and night variations. 

Influence of Diet. — The sugar output is increased by a carbohydrate 
diet, especially by dextrose and its polysaccharides. If a diabetic be fed 
levulose he will use it fairly well for a day or two and then excrete it as 
glucose. The starches of potato and oatmeal would seem to be very well 
borne and yet this relationship between diet and glycosuria is in some 
measure only apparent since the sugars differ much in the rate of their 
absorption and the ability of the body to store instead of to use them. 
For illustration, the " good " results observed of the potato and oatmeal 
" cures " were in part explained by the partial starvation of these badly 
fed patients and second by the ease with which these starches are digested 
by the external secretion of the pancreas which therefore is less fatigued 
and so better able to produce its internal secretion. In severe cases the 
variation in the sugar output depends in part on gastro-intestinal troubles 
so common in diabetes which may prevent the absorption of sugar. Muscle 
work, up to a certain point, decreases glycosuria, while psychical influences 
such as fright, mental strain ; or worry will increase it much and may even 
bring a latent case to light. Hence in the treatment of diabetics moderate 
physical work and a peaceful mind are important points. Mendel and 
Lusk 87 found that in completely diabetic dogs on a constant proteid-fat 
diet the ratio of glucose to nitrogen in the urine was constantly 3.65 : 1. 
But no man yet tested has been found completely diabetic; that is, all can 
burn some glucose and certainly one is never justified in exposing his patient 
to the injury which the above diet test would entail. It is interesting that 
Geyalin and DuBois' case (see page 199) for a considerable period have had 
D : N = 3.6 : 1, i.e., had Lusk's " fatal ratio " and yet improved. 

Intensity of Glycosuria. — On a rich carbohydrate diet the percent- 
age of glucose may reach, but rarely exceeds, 6 to 8%. Naunyn men- 
tions a case with 11%, while others mention a case with 20% of glucose 
in the urine. 

The effect of acute infections on a glycosuria is variable and often par- 
ticularly interesting. In pneumonia, for instance, a remarkable diminu- 
tion in the sugar output due to an increased tolerance to carbohydrates 
begins with the rise of temperature and is not clue to the diet. No satis- 
factory explanation has been given. On the other hand the sugar output 
may be increased during a fever or a glycosuria may begin with and continue 
after a febrile disease. This is the reason why so many diseases which have 
been contracted by patients with latent glycosuria have been reported as 
the " cause " of diabetes mellitus. 

87 Deutsches Arch. f. klin. Med., 1904, vol. lxxxi. 



202 CLINICAL DIAGNOSIS 

Chronic diseases such as tuberculosis of the lungs, diseases of the central 
nervous system, circulatory disturbances with albuminuria and nephritis 
tend to diminish the sugar output. As some diseases develop, and this is 
true of Blight's disease especially, the glycosuria progressively diminishes 
and finally disappears, hence they are reported as cured. This decrease 
in the glycosuria is due to a definite raising of the threshold point of the 
kidney for glucose and not to changes in the diet. Neither is it due to an 
increased inability of the kidneys to eliminate sugar since there is no increase 
in the hyperglycemia. An increase in the assimilation limit to glucose 
would seem to be a phenomenon of developing cachexia 

The output of sugar in each case of diabetes is subject to spontaneous 
and wide fluctuations which must be due to variations in tolerance. 

Severity and Tolerance. — A light case, according to Naunyn, is one 
which can eat daily 60 gms. of bread and remain sugar-free for a consider- 
able time. Those persons are said to have a " paradoxical tolerance " who 
can consume daily considerable carbohydrate with only a trace of sugar 
in the urine and yet who cannot get rid of that last trace. These are severe 
cases. The student should remember, first, that a pure proteid diet is not 
sugar-free and second that while severe cases may keep sugar-free for some 
time on a full proteid-fat diet, yet it is at great expense to the body. There 
is a constant tendency for a large glycosuria to increase and the greater the 
glucose output the less becomes the tolerance. The slight glycosurias tend 
to diminish. A patient's tolerance suffers more from the ingestion of a 
large amount of glucose at one time than from the same amount in divided 
portions. The reverse is also true that tolerance is increased more by a 
brief period during which the patient is quite sugar-free than by a much 
longer period of slight glycosuria, hence the value of the " hunger day " of 
Naunyn and of the fasting periods of Allen. If as the result of fasting a 
patient becomes sugar-free for even 24 hours he will on the following day 
be able to use without the production of a glycosuria an amount of bread 
which previously would have caused a marked rise in the output of glucose. 
That is, the question of tolerance would seem closely related to that of the 
fatigue of a gland. 

A case of " transitory diabetes " with acidosis is reported by Mann 88 
which lasted 16 days, then disappeared even though the patient consumed 
much sugar. 

Coma and Acidosis. — By " acidosis " Naunyn meant the accumula- 
tion in the body of those acid bodies which are formed in normal metabolism 
but which the healthy organism will rapidly eliminate, neutralize or 
further oxidize. The accumulation of these acid bodies produces an acid 
intoxication, or better expressed, an alkali starvation, and this it is believed 
is the explanation of diabetic coma. The laboratory evidence of acidosis 
includes the appearance in the urine of large amounts of acetone, diacetic 

88 Berl. klin. Wochenschr., 1904, No. 30. 



THE URINE: DIABETES MELLITUS 203 

acid, and, in severe cases, of /3-hydroxybutyric acid, probably the mother 
substance of the two, a great increase in the output of ammonia which serves 
as a base for some of these acid bodies, a retention by the tissues of any 
alkali administered and a decrease of the CO 2 tension of the blood. The 
production in the body of these acetone bodies is not characteristic of a 
diabetic disturbance of metabolism, since the urine of a fasting person 
and of a normal person on a sugar-free diet will in 3 or more days contain 
them all. But in the diabetic these remain unoxidized and they may 
excrete from 20 to 30 gms. of oxy butyric acid daily for years. When an 
acidosis once begins the tendency is for it to increase. It is increased greatly 
by a rigid diet. This is the reason why the introduction of the strict dietary 
treatment of diabetes, the carbohydrate-free diet consisting of much proteid 
and fat, was followed by a great increase in the frequency of diabetic coma. 
As coma develops there is usually a sudden increase of these acid bodies 
in the urine. A patient may excrete daily for even months 20 gms. of oxy- 
butyric acid, but if the output reaches 25 gms., on-coming coma should be 
feared (Herter). Any improvement in a patient in coma is followed by a 
greatly increased output of acid bodies, for it is not the acid in the urine 
which causes the trouble, but the acid which has not been excreted. Joslyn 
takes issue with this statement and feels that this increase is due to the 
administration of large amounts of alkali and is of no advantage to the 
body but rather is an injury. The presence of acidosis means that the 
case is a severe one, or at least that emaciation has begun, and yet such a 
patient may live for years. The amount of, or rather the danger from, 
these bodies has in the past been estimated by the ammonia output since 
the symptoms are caused especially by a withdrawal of the body alkalies 
which the ammonia protects. Any increase of ammonia output is said to 
mean the presence in the urine of at least 10 gms. of oxy butyric acid per 
day. A marked increase of ammonia indicates about 15 gms., while 4 gms. 
of ammonia per day indicates 16 gms. of the acid (Herter). Naunyn con- 
siders that the output of over 3 gms. of ammonia per day means danger 
of coma. Coma was the terminal event of 18 of Naunyn's 44 fatal cases, 
the most of them young persons from 21 to 30 years of age. 

Sellards ("Acidosis," 19 17) believes that for the determination of the 
degree of acidosis the titration of the blood is very inadequate, the methods 
of physical chemistry are of no clinical value, that Rountree's method is 
inaccurate, while the determination of urine ammonia is of no value since 
the ammonia may be increased in conditions without acidosis and may be 
normal or decreased when acidosis is present. He believes that Van Slyke's 
method of determining the carbon dioxide tension of the blood may be 
feasible. His own bicarbonate deficit test is, he believes, the most delicate 
of the tests which are specific for acidosis. By means of it one can detect 
very slight grades of experimental acidosis and moderate grades in diabetes 
before acetone appears or ammonia increases in the urine. 



204 CLINICAL DIAGNOSIS 

Sellards 1 Test. 89 — Normally 5 gms. of sodium bicarbonate administered 
by mouth will make the urine alkaline, but if the tissues have been depleted 
of their alkali, even 90 gms. may be injected intravenously without change 
of reaction of the urine. Such a retention is seen in diabetes mellitus, in 
the nephritis of cholera and lesser grades in chronic interstitial nephritis, 
in acute nephritis and in renal anteriosclerosis, but not in chronic paren- 
chymatous nephritis. 

Sellards sterilizes a 2% solution of sodium bicarbonate in an autoclave 
at a pressure of 10 poitnds for 30 minutes in an atmosphere of CO2 (gener- 
ated by adding sodium bicarbonate to the water supplying the autoclave) . 
The sterilized solution is kept in cork-stoppered bottles and is used not 
later than 2 or 3 days after sterilization. Small amounts may be ad- 
ministered by mouth but larger must be injected intra venously. The 
reaction of the urine is tested with litmus paper. If the urine is but 
slightly acid it should be boiled before it is tested. The urine is collected 
at 3 hour intervals. 

Frothingham 90 using several methods, studied groups of cases of dia- 
betes, syphilis, epilepsy, exophthalmic goiter, primary anemia, chronic 
nephritis, pneumonia, acute rheumatic fever, subacute nephritis, lung 
abscess, gastric ulcer, Addison's disease, cirrhosis of the liver, chronic 
cardiac disease, as well as single cases of 17 other conditions, found an 
acidosis in some cases of diabetes, chronic nephritis, pneumonia, acute 
articular rheumatism and in several acute febrile conditions in the miscel- 
laneous group. 

Acidosis and Uremia. — While a mild grade of acidosis may develop 
in cases of uncomplicated nephritis, yet this will be compensated for by the 
increased excretion of acids by the kidney until advanced uremia develops 
and then there may be accumulations of non-volatile acids in the blood 
sufficient to depress the tension of C0 2 of the alveolar air. This acidosis 
does not, however, run parallel to the nitrogen retention or the output of 
phenolsulphonephthalein . 91 

A sign of coma in diabetes always suggestive, though not conclusive, 
is the appearance in the urine of such large numbers of granular casts that 
they form a gross sediment. This may appear with the coma or give 
warning even 24 hours in advance (Kiilz's sign, page 272). 

Among the other urinary symptoms in diabetes is an increase in the 
output of the creatinin (even 2 gms. per day), the animal gum cf Landwehr 
(much), uric acid, phosphoric acid and sulphuric acid. In all of these 
conditions, however, the increase is due to the diet and not to the disease. 
Oxalic acid also is increased (even to 1.2 gms. per day), especially as the 
sugar disappears. 

89 Bull, of the Johns Hopkins Hosp., Oct., 19 12, vol. xxiii, p. 289. 
• 90 Arch, of Int. Med., Dec, 1916, vol. xviii, p. 1717. 
91 Peabody, Arch, of Int. Med., 1914, vol. 14, p. 236. 



THE URINE: DIABETES MELLITUS 205 

The urine of these patients often contains albumin. In some this is 
due to a complicating disease, but of Naunyn's cases of pure diabetes 32 
of 94 had albuminuria. Of these in 17 there was only occasionally a trace 
of albumin present, in 6 the trace was almost constant, while in 10 there 
was marked albiuninuria. Naunyn believes that an albuminuria may be 
an expression of the effect of a glycosuria on the kidney. On the other hand, 
it is interesting that as chronic nephritis develops a glycosuria gradually 
disappears. This explains many of the so-called " cures " of diabetes 
mellitus. In other cases a glycosuria and an albuminuria may alternate. 

Diabetes Insipidus. — Diabetes insipidus is a disease of long duration 
the chief symptom of which is a marked polyuria of normal urine of very 
low specific gravity. It is a rare condition. Futcher reported from the 
Johns Hopkins Hospital clinic but 4 cases, or 0.001% of admissions. It 
occurs particularly in young men, although Jacobi considered that fully 
25% of the cases are in children under 10 years of age. 

The cases may be grouped as primary or idiopathic, which have no 
known lesion, and the secondary or symptomatic. The latest and best 
theory for the primary cases is that they are due to an increase in the 
secretion from the posterior and intermediate lobes of the pituitary gland. 
The secondary cases are associated with tumors of the brain, cerebral 
trauma, cerebral hemorrhage, cerebral lues especially and basilar menin- 
gitis, which diseases may of course affect the pituitary gland. Marked 
polyuria may be associated with certain diseases of the abdominal 
viscera and of the spinal cord, it is an occasional symptom of the psy- 
choses, of hysteria, of epilepsy and of chorea. Some of these cases 
resemble closely diabetes insipidus, others suggest rather a primary poly- 
dipsia, since on limiting the intake of fluids the urine becomes almost 
normal in amount. 

Polyuria is the most conspicuous symptom of this disease. From 20 to 
40 (and in 1 case, 43) liters may be voided in 24 hours. The amount 
voided daily by two children was almost equal to their body weight. The 
urine is pale, watery in color, faintly acid in reaction, and with a specific 
gravity of from 1 .002 to 1 .005 or perhaps 1 .010. Failure to make the neces- 
sary temperature correction probably explains some of the impossible 
figures of specific gravity reported. In a few cases the specific gravity of 
the urine, although there was polyuria, is more nearfy normal (e.g., 6 liters 
and 1.017). The urine of these cases shows a remarkable fixation of the 
specific gravity, i.e., this remains almost constant in spite of all means 
taken to raise it (the ingestion of NaCl, etc.) . Albuminuria and cylindruria 
are absent, or if present mean a complicating condition. Sugar is also 
absent and yet there are cases of diabetes insipidus which later develop 
into diabetes mellitus and vice versa. Brackett's 92 case began suddenly 
as polyuria following mental shock, but 7 months lat er, just before death, 

92 Lancet, 1899, No. 25. 



206 CLINICAL DIAGNOSIS 

the specific gravity which had varied from 1.002 to 1.006 rose to 1.026 
and considerable sugar appeared. Some patients with diabetes insipidus 
would seem to void daily an amount of urine greater than the fluid intake 
(including the water content of the solid foods). The urine voided by one 
of Futcher's cases, for instance, exceeded the fluid intake by from 400 to 
6355 c.c. per day and yet this patient was carefully watched to prevent 
deception. The solids of the urine of a case of diabetes insipidus are in- 
creased because of the polyuria which " washes " these from the blood. 
This explains the remarkable polyphagia, as a result of which their urea 
output may reach 80 gms. or more per day. The output of sodium chloride 
and of phosphoric acid is either normal or slightly increased. Inosite is 
often present, probably washed out of the muscles by the unusual water 
flow due to the polydipsia since it can be found in the urine of normal 
persons after the ingestion of large amounts of water. 

Many believe that a condition of bradyuria exists in cases of diabetes 
insipidus ; that is, that fluid ingested is more slowly eliminated than normal 
but this is not constant. 

Glycuronic Acid, CHO(CHOH) 4 COOH— Glycuronic acid is an inter- 
mediate product of glucose metabolism which appears in the urine only 
when protected from further oxidation by conjunction with a suitable 
drug, as camphor, or with substances which arise in the body as indoxyl, 
skatoxyl, paracresol, phenol, or by combination with certain nitrogenous 
bases to form, e.g., uramidoglycuronic acid. The amount excreted, nor- 
mally less than 25 mgms. per 100 c.c. of urine, depends on the quantity of 
bodies present with which it can conjugate rather than on the amount of 
glycuronic acid formed. Free glycuronic acid crystallized with phenyl- 
hydrazine in beautiful needles whose melting point is 1 14° to 1 1 5 C. This 
acid, however, does not occur free in the urine, but must first be split from 
its paired compound by the addition, e.g., of acid and heat. The glycu- 
ronates will reduce copper, bismuth and silver, reducing copper as readily 
as it does glucose. They do not ferment. With HC1 and phloroglucin or 
orcin the free acid gives the same color tests as do the pentoses, including 
even their spectrum. The orcin reaction is the most convenient one to 
use (see page 185). It gives the furfurol test. While the free acid is dex- 
trorotatory the paired compounds, which alone appear in the urine, all are 
levorotatory and explain the levorotation of from 0.05 to 0.17 of normal 
urine. The amount excreted is much increased by the administration of 
camphor and of chloral hydrate and by all conditions which increase the 
production of the substances mentioned above with which this acid can 
pair. The clinical importance of the glycuronic acid compounds lies in 
the fact that they occur in normal urine in amounts sufficient to reduce 
copper after somewhat prolonged boiling and are levorotatory. If, there- 
fore, the reduction test for sugar is suggestive, the fermentation test nega- 
tive and the orcin test positive, they should be suspected. Since this acid 



THE URINE: DIABETES MELLITUS 207 

is a normal product of glucose oxidation, its elimination is increased in 
diabetes mellitus. Some, indeed, claim that in very mild cases of diabetes 
the urine contains increased amounts of this only and no glucose, but 
Edsall 93 showed that the excess of benzoyl esters in the urines of diabetics 
is not always due to an excess of glycuronic acid but rather to an increase 
of the unfermentable carbohydrates which are present in all intoxications 
and which may serve as a protective measure of the body to combat these 
intoxications, while Fisher has shown that the glycuronic acid is paired 
before any oxidation of the dextrose molecule has occurred. 

Alkaptonuria, — That alkaptonuria is a rare condition is evident from 
the fact that but 40 cases (29 of them men) had been reported up to 1902 
(Garrod) of which only 4 were in America (Futcher) , and that this number 
has not increased much since attention was called to the condition. The 
condition causes no symptoms and is discovered accidentally, by the 
mother, for instance, noticing that the napkins of the infant are darkly 
stained, or by an insurance company examining carefully an applicant 
whose urine seems to contain sugar. Urine supposed to be rich in pyro- 
catechin often proves to contain alkapton bodies. Alkaptonuria seems to 
be a congenital and life-long variant or freak of metabolism although some 
cases are intermittant, and Mittelbach's patient is confident that his 
followed an injury. The explanation given is that the body is unable to 
burn homogentisinic acid, an intermediate product of proteid catabolism. 94 
The condition is, therefore, comparable to glycosuria. Tyrosin, while the 
mother substance of some, cannot explain all of this acid in the urine. 

Alkaptonuria would seem to be a family disease, since 19 of 32 cases 
occurred in 7 families, in 1 of which were 4 cases. There is only 1 case 
indicating inheritance (Osier's case). 95 Garrod 96 finds that 60% of the 
cases are children of parents who are first cousins. Others think that it 
is due to an intestinal mycosis, a peculiar intestinal ferment, etc. The 
amount of reducing substance excreted in the urine varies from about 
3.2 to 6.9 gms. in 24 hours. It is of interest, said Garrod, that the output 
in each case is fairly constant and that each person excretes either the 
amount peculiar to himself or none. No traces, no gradual increases or 
decreases in the output are seen. Others find that the amount eliminated 
depends somewhat upon the diet ; that it is reduced a little by a vegetable 
diet and to about one-half by starvation. Mittelbach claimed that the maxi- 
mum excretion is from 1 to 3 hours after a heavy meal, which would indicate 
an intestinal origin, but Garrod found it to be from 4 to 8 hours after a 
meal, which would indicate a disturbance of tissue metabolism as the cause. 

The urine of a patient with alkaptonuria when fresh is very acid in 
reaction and normal in color, but it rapidly becomes dark, reddish-brown 

93 Univ. of Penn. Med. Bull., April, 1906. 

94 See Langstein and Meyer, Deutsches Arch. f. klin. Med., 1903, vol. lxxviii, p. 161. 

95 Lancet, Jan. 2, 1904. 

96 Lancet, Dec. 13, 1902. 



208 CLINICAL DIAGNOSIS 

and syrupy from oxidation, especially if the urine is made alkaline. It 
gives the copper test of glucose, but not Nylander's. AgN0 3 is reduced 
in the cold. It does not rotate polarized light, is not fermented and gives 
no crystals with phenylhydrazin. 

Of the alkapton bodies, 2 have been isolated; homogentisinic and uro- 
leucinic acids. There may be others. V. Jaksch includes the glycosuric 
acid of Marshall, which may, however, for the most part be homogenti- 
sinic acid. 

Homogentisinic acid C 6 H 3 (OH) 2 COOH, is the most important and 
in most cases the only alkapton body present. This is the substance which 
explains the characteristic reactions of the urine. Its mother substance 
would seem to be tyrosin, for this, if fed a patient in small doses, is excreted 
as this acid. 

To isolate homogentisinic acid the urine is made strongly acid with 
H2SO4 ( 1 to 12), 75 c.c. per 1 liter of urine. This is evaporated on a water 
bath to Ko its volume and shaken out 4 or 5 times with 3 volumes of ether. 
The ether is then distilled off, the residue dissolved in water (30 to 60 
volumes), filtered, the solution heated to boiling and precipitated with 
20% PbAc. This is quickly filtered while hot to separate the brown resin- 
ous precipitate. On standing the lead salts will slowly separate out. 
These are decomposed by H 2 S, and the nitrate carefully evaporated first 
on the water-bath and then in vacuo. The acid will crystallize out. Garrod 
recommends that the urine be heated to boiling and from 5 to 6 gms. of 
solid PbAc per 100 c.c. of urine added. When this is dissolved the urine 
is filtered and the filtrate allowed to stand 24 hours in a cool place. The 
lead crystals which separate out are ground fine, suspended in water, de- 
composed with H 2 S, filtered, evaporated first on the water-bath and then 
in vacuo to a syrup. 

Uroleucinic acid, C 6 H 3 (OH) 2 C 2 H 3 OHCOOH (?) also has been found 
in the urine of patients with alkaptonuria. Its reactions are very similar 
to those of homogentisinic acid from which it may be separated since it is 
precipitated by basic lead acetate. Garrod found none in the urine of 
some in whom years previously others had found it. 

Baumann's Quantitative Determination of Homogentisinic 
Acid. 97 — To 10 c.c. of the urine in a flask are added 10 c.c. of 3% ammonia. 
Then one adds at once several cubic centimeters of 0.1N AgN0 3 , shakes it 
a little and allows it to stand 5 minutes. Five drops of 10% CaCl 2 and 
10 drops of ammonium carbonate solution are then added. After shaking,, 
this is filtered. The brown-colored but entirely clear filtrate is tested with 
silver nitrate. If there at once appears a marked precipitate of reduced 
silver, the test is repeated, but a larger amount, even twice as much, of 
silver solution is added to the mixture of 10 c.c. of urine and 10 c.c. of 
ammonia. In this way one estimates approximately the amount of the 

97 Zeitschr. f. physiol. Chem., 1892, vol. xvi, p. 270. 



THE URINE: DIABETES MELLITUS 209 

silver solution necessary to oxidize the homogentisinic acid present. The 
end reaction is now determined by adding HC1. One is near this when on 
the addition of HC1 the deep brown fluid takes a light red color. The end 
reaction is reached when the filtrate from the silver precipitate acidified 
with dilute HC1 shows a slight precipitate of AgCl. One can determine 
this point very sharply by repeating the determination 4 or 6 times. If 
more than 8 c.c. of silver solution are necessary, on repeating the determina- 
tion 20 c.c. rather than 10 c.c. of ammonia should be used. 

One gram of the water-free homogentisinic acid is reduced with the 
above technic by a quantity of silver solution which contains 2.6 to 2.65 
gms. of silver; that is, 240 to 254 c.c. of the 0.1N silver solution. Hence 
1 c.c. of the 0.1N solution indicates 0.004124 gms. of the acid. The method 
has an average error of 6.1%. It is therefore only approximate. 

Fat in the Urine (Lipuria). — The urine of the normal person contains 
practically no free fat. The scum of calcium phosphate on the surface 
of some urines may closely resemble fat, and oil used in catheterization 
must of course be excluded. Lipuria may accompany a diet too rich 
in fat, fat medication, prolonged suppuration, phosphorus poisoning or 
diabetic lipemia. In urines containing a cell-rich exudate of pus-cells 
(as in pyonephrosis), epithelial cells and epithelial and fatty casts, con- 
siderable fat may be liberated by the disintegration of these formed ele- 
ments. In some cases, however, of ehyluria the urine is turbid with fat 
droplets. In these cases there are usually ruptured varicosed lymphatic 
vessels on the wall of bladder or in the kidney. Since lymph has a very 
variable fat content the term lymphuria has been used of cases in which 
the lymph which mixes with the urine is poor in fat and ehyluria if it is 
rich in fat. If, however, the lymph originates in the varicosed renal lym- 
phatics the term lymphuria should be used, but if it is the contents of the 
thoracic duct, ehyluria. Some of these cases are due to the blocking of the 
pelvic lymphatics or those of the pelvis of the kidney by the adult filaria 
worms. Other cases probably of nonparasitic origin are due to old abscesses 
which have established such anastcmosis. 

The urine in ehyluria often resembles milk or is opaque and red from 
the presence of blood also (hematochyluria) and sometimes coagulates 
into a jelly-like mass. 

In cases of ehyluria the fat content of the urine varies remarkably 
(from o to 1.4%) with, although not in direct proportion to, the fat content 
of the ingested food. 

To determine 98 the fat of the urine, 10 c.c. of the specimen to be ex- 
amined are mixed with about an equal amount of sand and evaporated 
to dryness on the water bath. The sand containing the urinary residue 
is then collected in filter papers and extracted with ether for 12 hours in a 
Soxhlet fat extractor. The ether extract is then transferred to a weighed 

98 Carter, Arch, of Int. Med., 1916, xviii, p. 541. 
14 



210 CLINICAL DIAGNOSIS 

dish and the ether evaporated by a current of air until the dish has assumed 
a constant weight. From the amount found in this way the amount of 
ether-soluble fat in the total specimen of urine is calculated. 

PROTEIDS IN THE URINE 

Tests for Albumin. — Urine to be tested for albumin should if possible 
be fresh but at least should always be clear. If it cannot be examined while 
fresh it should be protected from decomposition (see page 88), otherwise 
the rapid changes in its reaction may render the tests difficult. A turbid 
urine, the turbidity not due to bacteria, is best cleared by nitration through 
several thicknesses of paper. But when bacteria are present it is best 
filtered through infusorial earth, magnesia, or an asbestos filter although 
the infusorial earth may remove some of the albumin. A concentrated 
urine should always be diluted, since this will render albumin tests more 
sensitive rather than less so. 

Hallauer's " work emphasizes the importance of diluting the urine. If a normal 
urine be concentrated by heat to K its volume and then serum albumin added, the heat 
and acetic acid test will be sensitive, but the heat and nitric acid, Heller's and the potas- 
sium ferrocyanide tests all will be negative. If the urine be concentrated to % its volume 
and serum albumin added, none of these tests will show the albumin, and yet if we now 
dilute this specimen to its original volume, all of these tests will be positive. The potas- 
sium ferrocyanide test is the first to fail to show albumin, giving negative results after 
the specific gravity has reached about 1.030. It is the urea and the inorganic salts, 
especially phosphates, which inhibit the albumin tests. 

The specimen of urine to be examined should be chosen with care, 
since albumin may be present only during a few hours of the day. In 
doubtful cases therefore it is well to examine an afternoon specimen voided 
an hour or so after active exercise. This may be clearly positive for albumin 
although were that voiding mixed with the other albumin-free voidings 
of that same day the test might be negative. 

The Heat and Acetic Acid Test. — A clean test-tube is filled to within 
an inch of the top with filtered, clear urine. Holding the tube by its lower 
end, one heats its upper half to the boiling point and then holding the tube 
against a black background examines the boiled urine for a cloud. For 
careful work an alcohol flame is the best heat since the gas flame may deposit 
on the glass a faint film which suggests albumin. If turbidity appears 
in the urine it may be a cloud of albumin or one of calcium phosphate and 
calcium carbonate. To rule out the latter, a few drops of 5% acetic acid 
are added until the urine is distinctly acid. The cloud of phosphates and 
carbonates will promptly disappear, the carbonates with effervescence. 
Instead of acetic acid Hammarsten recommends from 1 to 3 drops of a 
25% HC1 per 10 c.c. of urine. After the addition of each drop of acid the 
boiling should be repeated. The acid will render an albumin cloud more 
distinct and more flocculent. 

99 Munch, med. Wchschr., 1903, p. 1539. 



THE URINE: PROTEIDS 211 

If the heat does not produce a cloud the acid should nevertheless be 
added, since the urine may not be acid enough to permit the albumin to 
coagulate. And, even though the boiled and acidified and again boiled 
urine remains perfectly clear, it may nevertheless contain a demonstrable 
amount of albumin which will appear as a distinct cloud if the tube be 
allowed to stand for about 15 minutes. It is the failure to observe this 
precaution which leads to most mistakes. It is said that some very acid 
albuminous urines are not clouded by boiling unless a drop of alkali be 
added. Acetic acid in excess does not produce soluble acid albumin. This 
is, we believe, the most reliable of all the routine tests for albumin and 
yet it may be improved by a slight modification. 100 To the urine is added 
% its volume of a saturated aqueous solution of sodium chloride and from 
3 to 5 drops of 50% acetic acid. One then proceeds in the manner described 
above. The sodium chloride renders the test more sensitive and holds all 
nucleo-albumin in solution. 

The more chronic the nephritis the whiter the albumin cloud. The 
more acute, the browner it is. 

Another coagulum which may appear in urine thus tested is the so-called 
" nucleo-albumin " or mucin, but this would also be precipitated in the 
cold by acetic acid (see page 220). One should therefore always make a 
control test by adding the same amount of acetic acid to a similar amount 
of unheated urine. The urine gives a better test for nucleo-albumin if 
diluted; this precipitate is soluble in an excess of acid. 

Resinous acids in the urine may lead to error if a great excess of acetic 
acid is added. This cloud is soluble in alcohol. This precipitate may be 
met with after the ingestion of certain resinous bodies : turpentine, benzoin, 
copaiba, balsam of Peru, tolu, cubebs, etc. ,1 

Heat and Nitric Acid. — This test differs from the above in that con- 
centrated HNO3 is used instead of dilute acetic acid. The HN0 3 should 
be used in fair excess since the danger is that too little rather than too much 
may be added. Hammerstein recommends to add 1 to 2 drops of 25% 
HNO3 for each 1 c.c. of urine. The urine should be boiled before and 
after each addition of acid. A flocculent precipitate obtained under these 
conditions indicates albumin, since the phosphate clouds are dissolved and 
nucleo-albumin is soluble in this excess of HN0 3 . A cloud which either 
appears or increases after the acidified and boiled urine is allowed to cool 
is of " albumose." Uric acid may precipitate when the urine cools but 
this cloud is granular and colored. To demonstrate a faint trace of albumin 
by this test the saturated chloride mixture should be added to urines 
of low specific value (see above). Since a very great excess of HN0 3 
may dissolve a mere trace of albumin, the acid should be added gradu- 
ally and the boiling repeated after each addition. In this test also the 
coagulation of a trace may not be evident until the specimen has stood 

100 See Hasting, Medical Record, July 7, 1906. 



212 



CLINICAL DIAGNOSIS 



for some time, then a coagulum would if present be found at the bottom 
of the tube. 

This test excludes the " albumin normally present," the " nucleo- 
albumin," and indicates the nature of "albumose" (Bence- Jones body) if 
present. The urates may deceive one if the urine is concentrated, but this 
precipitate is never flocculent ; the same is true of the resinous acids. Biliver- 
din and other pigments may cause a cloud but this is soluble in alcohol. 
The heat and acid tests are very delicate, indicating as they do 0.005 
gm. of albumin per 100 c.c. of urine. They 
should, however, always be confirmed and they 
do not always indicate the albumoses. 

Heller's Nitric Acid Test. — This is a con- 
tact test between urine and cold pure nitric acid. 
If albumin is present a line of precipitated acid 
albumin, insoluble in a fair excess (but soluble in a 
great excess) of the acid, will form at the plane of 
contact. This test has one advantage that no 
heat is necessary. Of all the mineral acids which 
have been used, nitric required the least acid per 
molecule of albumin to give an insoluble com- 
pound. This test is delicate, indicating, some 
say, 0.007% an d others, even 0.002% (Ham- 
marsten) of albumin. The best vessel in which 
to perform the contact test is a very large test- 
tube or a wineglass, or best of all a U-shaped tube 
like that pictured below (see Fig. 34). 

As usually applied one pours into the tube 
about 2 inches of urine and then, holding the tube 
as horizontal as possible, allows the concentrated 
nitric acid to run slowly, best from a pipet, down 
the bottom of the tube. The object is to underlay 
3 volumes of urine with 1 of HN0 3 and mix the fluids as little as possible. 
Some pour the urine in over the nitric acid. The nitric acid must be 
colorless, i.e., contain no nitrous acid, since the effervescence which 
this and the urea at the line of contact will produce would prevent 
the formation of the ring and so a faint trace of albumin would go 
unrecognized. The same would be true if much carbonate is present, 
as in an old urine. The ring may not appear for several minutes. To 
see the faint line indicating a trace of albumin one should hold the tube 
against a dark background. If the ring appears at the end of 3 minutes 
the albumin content is less than 0.003%. This precipitate of acid albumin 
will develop exactly at the plane of contact ; its thinness and density depend 
on the amount of albumin present and also on the skill with which the two 
fluids have been superimposed. 




Fig. 34. — Horismascope. A, 
the arm of the U-shaped tube 
with fine bore; B, bulb in which 
HNO3 is poured after the tubes 
are filled with urine; C, wide- 
bore arm for urine, with back- 
ground. 



THE URINE: PROTEIDS 213 

A red or reddish-violet ring at the plane of contact appears if an albumin- 
free urine is concentrated. This should not deceive one since it contains 
no precipitate. 

A ring of precipitated urates often appears if the urine is concentrated 
but this is always above the line of contact and separated from it by a 
layer of clear urine. It also is broader than the albumin ring, is less dis- 
tinct, it disappears on warming and will not appear if the test is repeated 
after the urine has been diluted with about 2 volumes of water, which dilu- 
tion would often improve the albumin test. Sometimes, although rarely, 
this precipitate is very dense, clouding the whole volume of urine. Since 
this will disappear on warming and reappear on cooling it has been mistaken 
for the Bence-Jones body and a hopeless prognosis given. 

Nucleo-albumin. — The body called often nucleo-albumin may give 
an opalescent ring 0.5 to 1 cm. above the line of contact and sometimes 
extending down to it. But this will disappear or at least move up (since 
the ring below disappears and a new ring forms above) if the tube is slightly 
shaken, since it is soluble in nitric acid. This ring may appear after the 
tube has stood for some minutes, it is faint and does not resemble the 
albumin ring much in appearance. If the urine is diluted and the test 
repeated, this precipitate will appear more rapidly and be more distinct. 

Resinous Acids. — Heller's test applied to a urine containing resinous 
acids may produce a whitish ring above the line of contact of acid and urine 
which partly clears on warming. Since this precipitate is soluble in ether, 
to exclude the resinous acids one may pipet off the turbid layer of urine 
and mix it with a great excess of ether to prevent an emulsion. Or, one 
adds 2 or 3 drops of HO to from 8 to 10 c.c. of the cold urine which 
will precipitate these acids. If then one adds more HC1 and heats the 
urine, a red color will result. To remove these resinous acids from the 
urine one makes it strongly acid with acetic acid and extracts it with ether. 

If the " albumoses " (Bence-Jones body) are present, Heller's test will 
give at the line of contact a very heavy ring which disappears on warming 
and reappears on cooling. 

The bile acids will give a precipitate if present in concentrated urines. 

If the urine is rich in urea Heller's test may produce at the line of con- 
tact a solid crust of urea nitrate which is so solid and so definitely crystalline 
that it should never deceive one. 

Hammarsten recommends that as a routine all urines to be examined by 
this test be diluted to a specific gravity of 1.005 since then all of the above 
disturbing bodies will be excluded except albumose and nucleo-albumin. 

This test for albumin should always be confirmed by one of the other 
tests. Many workers recommend that this be the one used first. 

Potassium Ferrocyanide and Acetic Acid Test. — The urine is 
made quite acid with a few drops of acetic acid and then 5% K 4 FeCN 6 is 
added drop by drop. The presence of albumin will be indicated by a cloud 



214 CLINICAL DIAGNOSIS 

or flaky precipitate. When this ceases to increase no more of the reagent 
should be added. In the hands of an expert this test is more accurate 
than is Heller's. Success in its use depends on the proportions of the 
reagents used and on the amount of salts present in the urine. The test is 
particularly valuable in quantitative work to determine, e.g., whether or 
not all of the albumin has been removed from a solution. 

The albumoses also are precipitated by this test as is also " Nucleo- 
albumin," but the latter is also by acetic acid alone. 

The urine should not be tested while hot nor should any reagent used 
contain iron (as Kieselguhr) for this would give an abundant precipitate. 

Tanret's Test. — Tanret's reagent contains 1.35 gms. of HgCl 2 dis- 
solved in as little water as possible together with 3.32 gms. of KI. To 
this solution are then added 50 c.c. of water and finally 20 c.c. of glacial 
acetic acid. This reagent is added to the urine drop by drop until the cloud 
of albumin just begins to appear. This test is exceedingly delicate. It 
indicates also "nucleo-albumin,'' "peptone" (soluble on warming), alka- 
loids and the albumoses. We have seen this test used in French clinics 
with the most satisfactory results. 

Spiegler's Test. — Spiegler's reagent as modified by Jolles contains 
HgCl 2 , 10 gms.; succinic acid, 20 gms.; NaCl, 10 gms.; and water 500 c.c. 
The specific gravity of this fluid should be well above 1.060 in order that 
it may be used as a contact test. 

This is the most delicate test of all for albumin. The urine is first 
filtered, rendered acid by a few drops of acetic acid to hold the carbonates 
in solution and to precipitate any nucleo-albumin, which if present should 
be filtered off (otherwise it would be precipitated by the reagent). The 
urine is then superimposed on this reagent (see page 212). If albumin 
is present a very sharp definite ring will appear at the line of contact. 
It is claimed that this test will be positive for 1 part of albumin in 
150,000 to 350,000 of urine. It is positive also for the albumoses, but not 
for deutero-albumose. 

Various other tests have been proposed. For convenience sake some 
add to a test-tube of urine a piece of solid metaphosphoric acid or of picric 
acid the size of a pea. A long list of very delicate tests have been recom- 
mended, but there is little to recommend then since it is granted that all 
normal urines contain a little serum albumin. The important thing is 
that a worker use 2 tests which control each other well, understand the 
shortcomings of each and be experienced in their use. 

The order of delicacy of the tests mentioned above is: Spiegler's, Tan- 
ret's, heat and acid; K 4 FeCN 6 ; Heller's, picric acid, etc. This order, given 
by Huppert, is not accepted by some, who claim that Heller's test properly 
performed in a wineglass gives more delicate results even than the heat and 
acid-test. Senator recommends Heller's test, since it shows albumose. He 
advised against the heat test, since by its use traces of albumin and rather 



THE URINE: PROTEIDS 215 

large amounts of albumose are so often lost. We would recommend for 
routine work the heat and acetic acid-test after the concentrated salt solu- 
tion has been added to the urine and that this be controlled by Heller's test. 

Quantitative Determination of Albumin. — Scherer's Method Modi- 
fied by Cohnheim. — To a carefully measured amount of urine (about 
500 c.c.) is added Xo its volume of a saturated solution of sodium chloride 
and then it is filtered. About 5 c.c. of this urine is then boiled in a test- 
tube and filtered. The nitrate is tested for albumin with acetic acid and 
potassium ferrocyanide. If this test is negative for albumin then the acidity 
of the whole volume of urine is correct and one should proceed at once with 
the determination. If, however, this test shows a trace of albumin in the 
filtrate, then 2 or 3 drops of 50% acetic acid (and only 1 if but a trace was 
present) are added to the whole volume of urine. This is well stirred and 
then another 5 c.c. are removed and tested as before. The acidity of the 
whole volume of urine thus repeatedly increased (or decreased if necessary 
by a drop of strong NaOH solution) and samples tested until 1 is made 
albumin-free by heat. Two carefully measured quantities (see below) of 
the urine, 1 as a control, are now heated in beakers, first on a water-bath 
and then over a free flame, until the precipitate is flocculent and the super- 
natant fluid clear, and are then filtered through a weighed filter. 

The quantity of urine to use in the analysis should be such that the 
weight of the dried albumin precipitate will lie between 0.1 and 0.3 gms. 
If below 0.1 gm., the limit of error is too great; if above 0.3 gm. it is prac- 
tically impossible to dry the precipitate to constant weight. 

Urine which contains much albumin should be accurately diluted with 
salt solution. If the urine is very rich in albumin, the best method is to 
pour drop by drop a small, accurately measured amount of the urine into 
a beaker of boiling, half-saturated salt solution. 

When coagulation is completed by boiling the urine over the free flame, 
the urine is filtered through a dried and weighed filter paper and the pre- 
cipitate washed free from chlorine (a few drops of filtrate are tested at 
frequent intervals with AGNO 3 solution) with hot water, then with alcohol 
and finally with ether. The paper containing the precipitate is then 
placed in a weighed glass (with accurately fitting cover) and dried in an 
oven, the temperature of which is held at no° C. The glass in the oven 
should rest on a sheet of asbestos and the bulb of the thermometer of the 
oven should hang at the level of the glass. At intervals of about one hour 
the glass is removed from the oven with a pair of tongs and cooled in a 
desiccator. The cover is then inserted tightly, and the glass weighed. 
This is repeated until constant weight is reached. Even with most careful 
work the controls may differ by even 1%. 

Salkowski 101 advised that if there is an unusual amount in the 
urine a small, accurately measured volume of the specimen be mixed 

101 Berl. klin. Wchschr., March 3, 1902. 



216 



CLINICAL DIAGNOSIS 



with from 10 to 20 volumes of 05% alcohol and this brought to the boiling 
point on the water-bath. It is then cooled, the supernatant fluid decanted, 
the precipitate washed with hot water, filtered through a weighed filter, 
washed as above, then placed in a weighed platinum crucible and brought 
in an oven to a constant weight. It is finally burned and the weight of the 
ash determined and subtracted from that of the precipitate. 
Esbach's Tubes. — An Esbach tube (see Fig. 35) is filled to 
the mark U with urine and to the mark R with the reagent. 
The tube is corked, reversed slowly 12 times and then left stand- 
ing in a tube rack at constant temperature for just 24 hours. 
At the end of that time the height of the precipitate is noted. 
The figures on the scale indicate the number of grams of albumin 
per 1 liter of urine. 

Esbach's reagent (picric acid, 10 gms. ; citric acid, 20 gms.; 
water, sufficient to make 1 liter) has not given satisfactory 
R results and has been replaced by Tsuchiya's reagent. 

Phosphotungstic acid, 1.5 gm. 
Concentrated HC1, 5 c.c. 

Ethyl alcohol, q.s. ad 100 c.c. 

This method is quite accurate enough for clinical work 102 
provided Tsuchiya's reagent be used. 

An approximate estimation of the amount of albumin pres- 
ent can be based on Heller's test, made in a " Collamore 
wineglass " half filled with urine, then underlaid with approx- 
imately % its volume of nitric acid. By " slightest possible 
trace" is meant the smallest amount of precipitate which can 
be detected as a hazy ring under most favorable conditions 
(black background, etc.); " very slight trace " means slightly 
more; a "slight trace" can be seen without a background and 
also from above, although the bottom of the glass is distinctly 
seen; a "large trace" (about 0.1%) is a ring clearly seen but not 
flocculent, quite dense but not opaque when seen from above; 
the bottom of the glass cannot be seen through the ring made by 
0.15% although a faint ray of light is transmitted; 0.25% of 
albumin gives a zone quite flocculent from the side and quite 
opaque from above; 0.5% and above give a ring which is very 
dense and flocculent. Above this one cannot go by this method. The 
width of the ring is not so important (condensed from Ogden, " Clinical 
Examination of the Urine") . This method determines at the same time the 
" nucleo-albumin " and resinous acids present. 

Centrifuge Method. — Purdy recommended the use of graduated 
centrifuge tubes in which are mixed 10 c.c. of filtered urine, 3.5 c.c. of 10% 
K 4 FeCN 6 and 1.5 c.c. of acetic acid. The urine is then centrifugalized at a 
102 See Mattice, Arch. Int. Med., March, 1910, vol. v, p. 313. 



•U 



Fig. 35-— 
Esbach's 

albumi- 
nomet er . 



THE URINE: PROTEIDS PRESENT 217 

uniform speed of 1500 revolutions per minute in a centrifuge, the arm of 
which is of such length that the distance from the center of rotation to the 
tip of the tube is 7% inches. Each tube is centrifugalized 3 times, 5 minutes 
each time ; Xo c.c. volume of precipitate indicates %o% by weight of albumin. 
He gave a table with the equivalents of the readings. This test, satisfactory 
as it may seem, has not given very good results in our hands, although 
better than has the Esbach tube. It is an interesting fact that 2 of the 
makers of the " Purdy centrifuge " were unable to supply us with an arm 
which conformed to his specifications as regards length (hence they had to 
be made to order) or with graduated tubes with the sharp point as he 
represents them. We have found it is no. easy matter to keep a centrifuge 
running uniformly at the rate specified unless one carefully watches the 
taxometer and yet the exact time and speed of rotation are of great im- 
portance. 

Roberts and Stolnikow's Method. — This method is based on the 
observation that if with Heller's test a ring appears in from 2% to 3 minutes 
after the test is made there is an albuminuria of 0.003%. Different dilu- 
tions of the urine are therefore tested until there is one obtained in which 
the ring appears in 3 minutes. The test should be performed very care- 
fully. The sides of the tube should not be wet with the nitric acid and the 
urine should be added slowly from a pipet. 

It is often necessary to remove all albumin from a urine before under- 
taking other quantitative work. Usually it is sufficient to add acetic acid 
and to boil until the nitrate is clear (see page 215), and then restore the urine 
to its original volume. Hofmeister's method is more accurate. He adds 
to the urine an excess (10 c.c.) of a 40% solution of sodium acetate and 
concentrated Fe 2 Cl 6 until the specimen is of a red color. The urine is then 
neutralized or made very faintly acid, and boiled. The precipitate of basic 
ferric acetate which forms will carry down with it all of the albumin and 
leave an albumin- and iron-free solution which niters beautifully. This 
method cannot be used if glucose is present since then some ferric oxide 
will remain in solution. 

PROTEIDS PRESENT IN THE URINE 

By albuminuria is meant the presence in the urine of a coaguable 
protein. A true albuminuria is one which is due to disturbance of the 
cortex of the kidney (for false albuminuria, see page 224). The proteins 
usually present are serum albumin, " serum globulin, " and the so-called 
' ' nucleo-albumin . 

The albumin quotient is obtained by dividing the amount of serum 
albumin by the amount of " serum globulin " present (Hoffman). This 
quotient varies considerably in different cases, and in the same cases at 
different times. In some cases serum albumin alone has been found. 
This was true of 1 case of cancer of the stomach, and during limited periods 



218 CLINICAL DIAGNOSIS 

of certain cases of nephritis. Globulin alone was found in i case of acute 
nephritis, in i case during the puerperium and in i case of leukemia. (For 
" nucleo-albumin," see page 219.) 

Serum albumin is present in normal urine in amounts reaching even 
from 22 to 78 mgms. per 1 liter (Morner). Serum albumin is soluble in 
water and is coagulated by heat, if in acid solution, at a temperature vary- 
ing from 5 6° to 8i° C., depending on the amount of urea- and of salts 
(especially of phosphates) present and lastly on its own concentration, 
It is coagulated by alcohol, which coagulum if produced by absolute alcohol 
is soluble in water unless it has been in contact with the alcohol for a long 
time. The coagulum produced by weaker alcohol is more insoluble than 
that produced by stronger. The solubility of this coagulum should always 
be borne in mind when working quantitatively with albumin. Serum 
albumin is levorotatory, [a]D= —62. 6°. Serum albumin unites with an 
alkali to produce a soluble body which, when combined with a base, 
forms an albuminate much less soluble in water than is albumin itself 
and which may explain the spontaneous precipitate of albumin in some 
concentrated urines. 

The acid albumin produced by mineral acids is quite insoluble except 
in large excess of the acid while that produced by acetic acid is soluble in 
a very slight excess of the acid. 

Serum globulin is a term under which are included several quite differ- 
ent proteins, among them pseudoglobulin, euglobulin, and nbrinoglobulin 
(Hofmeister) , all of which exist in the blood-plasma and the reactions of 
which are rather different. Euglobulin and nbrinoglobulin (fibrinogen) are 
probably always present in the normal urine. It is they, slightly increased, 
which explain the mildest forms of albuminuria (the so-called physiological 
cases), while in severer cases serum albumin also is increased. 

These globulins may be separated by fractional precipitation by a 
saturated (NH 4 )2S0 4 solution. The limits of precipitation expressed in 
the number of cubic centimeters of the ammonium sulphate solution in the 
ioc.c. mixture are: pseudoglobulin 3.4 to 4.6; euglobulin 2.8 and 3.3; 
nbrinoglobulin, 2.2 to 2.9. 

Pseudoglobulin is not precipitated by acetic acid alone. 

Euglobulin occurs in almost all exudates and transudates and in 
many urines, perhaps in all. It can sometimes be precipitated by acetic 
acid in the undiluted urine, but usually one must dilute this with 2 or 3 
volumes of water. The acetic acid must be carefully added since the 
precipitate is easily soluble in excess. 

Serum globulin, including under this term all the above globulins, 
is present in the urine in cases of albuminuria in amounts varying from 
8 to 60% of the total proteid present; very rarely a trace only is present. 
In the blood its ration to serum albumin is as 1 : 1.5. Its greater relative 
ratio over the albumin in the urine cannot always be explained by its greater 



THE URINE: PROTEIDS PRESENT 219 

diffusibility, since euglobulin, which is constantly present, is little diffusible. 
The variations in the albumin quotient (see page 217) in nephritis are due 
to variations in the amount of globulin. Oswald considered that in the 
mildest form of albuminuria euglobulin alone is present in pathological 
amounts and that this is precipitated by acetic acid in the cold. This body 
is excreted in largest amounts in parenchymatous renal lesions. As a case of 
nephritis improves the relative amount of globulin in the urine diminishes, 
to increase with each acute exacerbation. In cases of contracted kidney and 
in nephritis with chronic passive nephritis it may be very low. It is fairly 
low in the albuminuria of pneumonia, but high in that of typhoid tever. 

The globulins are insoluble in distilled water. In the urine they are 
held in solution by the salts present. If, therefore, to a beaker of distilled 
water a drop or so of urine containing much globulin be added, a distinct 
cloud will appear. They may be detected also by diluting the urine till 
its specific gravity is about 1.002 and then adding 1 drop of acetic acid. 

To isolate the globulins the phosphates are removed by rendering the 
urine alkaline with ammonia and filtering. An equal volume of cool 
saturated ammonium sulphate is added to this filtrate, which will precipi- 
tate the globulins perfectly in neutral solution. The mixture is allowed to 
stand 1 hour and filtered. The precipitate which contains also albumose 
and " nucleo-albumin " is washed with half -saturated ammonium sulphate 
until the nitrate is albumin-free. Ammonium urate also may be precipi- 
tated in time and is to be avoided by working fairly rapidly. Serum 
albumin will not be precipitated until the point of total saturation with the 
sulphate is reached. The precipitate on the filter is now dissolved in a little 
water and heated on a water-bath which will coagulate the globulin, fibrino- 
gen and albumose. This is filtered and the precipitate washed with water 
and digested on a water-bath with 1% soda. This solution is then filtered 
and neutralized carefully with acetic acid. The precipitate which now 
falls consists of globulin and fibrinogen, but not albumose. 

To determine globulin quantitatively the filtered urine is first rendered 
neutral with ammonia and is then mixed with an equal volume of saturated 
ammonium sulphate solution. The mixture, well stirred, is allowed to 
stand for several hours. It is then filtered through a dried and weighed 
filter and the precipitate washed with half -saturated ammonium sulphate 
until chlorine-free. This filtration is a tedious process. The funnel con- 
taining the precipitate on the paper is then placed in a thermostat and 
dried for % hour at no° C. The ammonium sulphate is now washed out 
with hot water, the precipitate dried with alcohol, then with ether and 
dried at no° C. to constant weight (see page 215). In this deterariination 
also the amount of urine used should be such that the weight of the protein 
precipitate will not exceed 0.3 gm. 

Euglobulin, Nucleo-albumin, Mucin, Morner's Body. — If one 
adds to cold urine, and especially if this be well diluted, a few drops of 



220 CLINICAL DIAGNOSIS 

dilute acetic acid, there cften appears an opalescence or even a true precip- 
itate, which is difficultly soluble in an excess of the acid. When using 
Heller's test also one often sees a ring, not at the line of separation, but 
from 0.5 to 1 cm. above it. If serum albumin is present one may see both 
rings, and indeed the upper, the " nucleo-albumin " ring, is best seen in 
the urine of nephritis. The resinous acids and the urates should be excluded 
(see page 213). This proteid, commonly called nucleo-albumin, which 
explains these 2 phenomena, coagulates at about 5 6° C. It could be demon- 
strated in probably every normal urine if the salts be first removed by chassis . 

It was this body which led to the belief that at least one true proteid 
is a constituent of normal urine. Two other and contrary views have been 
held, one that this body is mucus, the other that it is true nucleo-albumin. 
If it were either of these, the condition would not be a true albuminuria. 
At present most believe that this substance is globulin or a compound of 
serum albumin and that a true albuminuria is normally present. 

A definite precipitate (indicating a definite increase of the protein) on 
the addition of acetic acid to the cold urine is seen in many conditions. 
Excluding the vesical cases, in which the substance is probably mucus, 
it is increased in the new-born ; in adults after severe exercise ; in nephritis ; 
in various acute diseases, especially those affecting the kidneys; in fevers, 
especially pneumonia and typhoid (also erysipelas, pleurisy, relapsing fever, 
meningitis, etc). Its increase in leukemia, reported first by Fr. Muller, 
gave rise to the idea that it was derived from the nuclei of the leucocytes. 

Oberma}^er found it in 32 cases of jaundice in amounts varying with the 
intensity of the jaundice; in scarlet fever in small amounts, in diphtheria 
in the greatest amounts of all ; after poisons affecting the kidneys (pyrogallic 
acid, corrosive sublimate, etc.) ; in acute yellow atrophy and after compres- 
sion of the thorax. 

In true nephritis the increase of this proteid may precede the true 
albuminuria and also may succeed it as the case improves. Madson con- 
siders it evidence of the earliest possible irritation of the kidneys. It may 
persist during the intermissions of an albuminuria. Euglobulin and 
fibrinogen are said to be the chief proteids in the urine in amyloid disease. 

In orthostatic albuminuria this may be the only proteid in the urine, 
or it may be accompanied by serum albumin and pseudoglobulin. In 
febrile albuminuria this may exceed in amount the serum albumin. It is 
present in traces in chronic interstitial nephritis. When the blood-supply 
of the kidney of animals is partly cut off, this body may be excreted in 
abundance, sometimes alone and sometimes with albumin. The same is 
true in partial suffocation of the organism. 

Pure mucus is present in the normal urine in traces (4.5 gms. in 260 
liters) and in 2 forms, an insoluble form which gives the nubecula and a 
soluble portion precipitated by acetic acid, which constitutes but a very 
small fraction of the whole. One would expect to find some mucus in the 



THE URINE: PROTEIDS PRESENT 221 

urine since the urinary passages are lined with mucous membrane the secre- 
tion of which will be washed off by the urine. This mucus is greatly in- 
creased in catarrhal conditions of the urinary tract. It is soluble in am- 
monia, is precipitated by acetic acid and is soluble in excess of this acid. 
From it a reducing body may be split off. This mucus does not contain 
nuclein nor chondroitin and the precipitate with acetic acid lacks the slimy 
character of true mucus thus treated. For this reason it has been called 
a " mucoid " body. It resembles the ovomucoid of the hen's egg (Morner). 
In the mine of a case of prostatitis we were able to obtain by precipitation 
with acetic acid 0.066 gm. of this substance per 100 c.c. of urine. 

To determine the mucus quantitatively the urine is precipitated carefully 
with acetic acid and repeatedly filtered through a weighed filter till the 
filtrate is clear. The precipitate is then washed with cold water acidulated 
with acetic acid, dried and weighed. Another method, much more rapid 
than the preceding but giving slightly lower results, is the following: A 
small amount (0.5 gm.) of infusorial earth dried at no° C. to constant 
weight, is mixed with the urine after the mucus has been precipitated. 
Then the urine is filtered, the precipitate dried and weighed. Then the 
weight of paper and of the Kieselguhr are subtracted. 

There are on record interesting and rare cases of true mucinuria which 
are analogous to mucous colitis and to fibrinous bronchitis. In these cases 
mucus casts 1 to 10 cm. long and 3 to 4 mm. thick may be voided with the 
urine. Such was v. Jaksch's case of "ureteritis membranacea" in which a 
spiral cast of the ureter consisting of mucus and fibrin was voided; Frank's 
case which he named " pyelitis product iva " passed a cast of the pelvis 
and upper ureter. Four cases only of this condition are on record. In 
these cases the symptoms of the passage of the casts resembled those of 
renal colic 

Nuc leo-albumin. — From a study of the urine of jaundiced patients 
Obermayer decided that the proteid which in that condition is precipitated 
by acetic acid is true nucleo-albumin and assumes that all precipitates by 
acetic acid in the cold are the same. True nucleo-albumin may be present 
in the urine, but this is not the body which usually goes under that name 
and its presence is never normal. It is said that its source is the broken- 
down epithelial cells of urinary tubules, as in acute nephritis, in which 
condition this proteid is most often present and in greatest amounts, after 
the ingestion of poisons which affect the kidneys, in disturbances of the 
renal circulation and finally jaundice, in which case its source is the bile 
itself. Nucleo-albumin is said by some to be a constant constituent of the 
blood, and it is possible that a certain amount of this may reach the urine. 
True nucleo-albumin, it is said, is found in the urine in cases of catarrhal 
inflammation of the urinary tract with desquamation of the superficial 
cells of the mucosa, as in cystitis or pyelitis. In the case of women the 
genital tract is to be excluded as the source. 



222 CLINICAL DIAGNOSIS 

It is apparent from the above list that nucleo-albuminuria is said to 
occur in conditions in which one might expect it, and yet one does not find 
it where nucleo-albumin should be present in the urine in largest amounts, 
as in urines containing abundant pus and epithelial cells. Here it is often 
hard to get any precipitate at all on adding acetic acid. 

It is very clear to one reading reports of cases of so-called nucleo- 
albuminuria that seldom has the crucial test, the proof that the substance 
in question contains phosphorus, been applied. (It also would of course 
be difficult to prove this, since it would be very hard to exclude a contamin- 
ation with the inorganic phosphorus of the urine.) 

Again, the " salting out " points with ammonium sulphate of the so- 
called nucleo-albumin of the urine do not quite agree with those of true 
nucleo-albumin obtained by the breaking down of tissue, which are 103 
minimal o.i to 0.8, maximal, 1.6 to 2.2. 

To prove that a body is nucleo-albumin it would be necessary to show 
that it is insoluble in acetic acid, is precipitated by MgS0 4 , that when 
boiled with dilute mineral acids it gives off no reducing substance and that 
peptic digestion gives nuclein and phosphorus. The last 2 tests it is almost 
impossible to apply to the urine. 

Morner's Body. — Much light was thrown on the subject of nucleo- 
albuminuria by Morner 104 who proved that most of the so-called nucleo- 
albumin of the urine is a compound of true serum albumin with an albumin- 
precipitating body which is formed when acetic acid is added to the urine. 
By dialyzing large amounts of urine, adding 1 to 2 parts per thousand of 
acetic acid, and then shaking out the residue with chloroform, Morner 
obtained a precipitate averaging 41 mgms. (22 to 78 mgms.) per liter of 
urine which resembled nucleo-albumin. This proved to be a combination 
of serum albumin and certain other bodies, among them chondroitin- 
sulphuric acid, which was always found and in the largest amounts, nu- 
cleinic acid, which is sometimes present in traces, and taurocholic acid, 
which is often present in traces but which in the case of jaundiced urine 
may exceed in amount the other 2. The fact that 3 different compounds 
of serum albumin may arise and in varying proportions explains the lack 
of uniformity in the properties of the precipitates which appear when acetic 
acid is added to the urine. This combination of albumin and these acids 
probably occurs after the acetic acid is added. If after removing this 
precipitate a little albumin is added to the urine a second precipitate will 
appear, amounting to about 54 mgms. per liter, which shows that the 
albumin-precipitating bodies are in excess of the albumin. The larger the 
proportion of serum albumin in this combination the more will the com- 
pound react like serum albumin, but the greater the relative predominance 
of these precipitating bodies, the more will it resemble nucleo-albumin. 

103 Matsumoto, Deutsch. Arch. f. klin. Med., 1903, vol. lxxv, p. 398. 

104 Skand. Arch. f. Phys., vol. vi, p. 332, 1895. 



THE URINE: PROTEIDS PRESENT 223 

It is this compound of albumin which explains the statements based on 
common experience, that " a true albuminuria is sometimes preceded by 
the excretion of a body precipitated by acetic acid " ; and that " the excre- 
tion of mucus may precede or succeed an albuminuria." It explains also 
the belief of recent years that this so-called physiological albuminuria was 
merely a nucleo-albuminuria. 

Morner used the following method of isolation: The salts of a large 
volume of urine are removed by dialysis and acetic acid then added, 2 c.c, 
per liter. The precipitate formed is then dissolved in a little water and 
again precipitated with acetic acid. A little is then heated for a long time 
on the water- bath with 5% HC1. If sulphuric acid is present but no reduc- 
ing body can be demonstrated chondroitin-sulphuric acid is probably 
present; if the reducing body can be demonstrated, but not sulphuric acid, 
the precipitate was probably mucus. If no sulphuric acid and no reducing 
body can be demonstrated, the precipitate should then be digested with 
pepsin and the products examined for organic phosphorus. If this phos- 
phorus is present the nuclein bases should be tested for. 

This explanation of Morner, satisfactory as it would seem and evidently 
based on very careful work, has received but little confirmation. Stahelin 105 
in 1 case of jaundice failed to find any of the " albumin-precipitating bodies " 
and thought the precipitate on adding acetic acid resembled the globulin, 
a view held by Fr. Muller in 1885 ; also in the very heavy acetic acid precipi- 
tate of the urine of a case of pneumonia no phosphorus could be detected. 
Matsumoto found this substance to consist chiefly of fibrinogen and euglo- 
bulin (see page 218). Oswald 106 studied this precipitate in the urines of 
cases of cyclic albuminuria and nephritis, and he also decided it to be 
euglobulin and a trace of fibrinogen. These proteins occur in the blood, 
but cannot be demonstrated in that fluid by the addition of acetic acid, 
since the salt content there is too low. 

It is to be noted that most observers have worked with smaller amounts 
of urine than Morner; again, that it is not proven that the limits of precipi- 
tation with saturated ammonium sulphate are sufficient for the recognition 
of a protein. In conclusion, it is clear that, however, the present conflict 
between Morner and the Hofmeister school may be settled, both agree 
that there is a constant normal physiological albuminuria. 107 

The nucleohiston of Lilienfeld is a body arising from the breaking 
down of leucocytes. It is precipitated by acetic acid and has a high 
phosphorus content. It has been found in large amounts in the urine of 
leukemic patients, although its appearance there is not due entirely to the 
breaking down of white cells. 

To demonstrate nucleo-histon in the urine all coagulable albumin is 
first removed. T hen all other proteids are precipitated with alcohol, the 

105 Munch, med. Wochenschr., 1902, p. 1413. 

106 Zeitschr. f. d. gesamt. Biochem., Bd. v., 1904. 

107 See also Calco., Zeitschr. f. klin. Med., 1904, vol. li. 



224 CLINICAL DIAGNOSIS 

precipitate washed in hot alcohol, then dissolved in boiling water, cooled, 
acidified with HC1, let stand and the uric acid precipitate filtered off. To 
the filtrate is then added ammonia, the resulting precipitate is collected 
on a small filter and washed with ammonia till the wash-water gives no 
biuret reaction. The precipitate is then dissolved in acetic acid and tested 
for hist on. This will give the biuret reaction, is coagulated by heat and 
this coagulum is soluble in mineral acids. 108 

Fibrin uria. — Fibrinogen or fibrinoglobulin is found in the urine rarely 
in demonstrable amounts. Its chemical reactions are those of globulin 
but its presence is indicated by the appearance of spontaneous coagulation 
if the urine is left standing. Excluding those cases in which there is con- 
siderable blood in the urine, fibrinuria is rare. It occurs in chyluria and 
rarely in nephritis. In some of these cases the urine clots at once after it 
is voided. The clot is sometimes firm and in other cases gelatinous. We 
have seen but i clear case, a woman admitted during the last hours of her 
life with what was evidently subacute parenchymatous nephritis. Only 
about 5 c.c. of urine could be obtained. This was cloudy, yellow in color 
and after standing for a few minutes clotted to a solid coagulum. In 
other cases reported the urine clots before it is voided and casts of the 
pelvis of the kidney or bladder are passed. In severe inflammation of the 
bladder, ureter, or pelvis of the kidney, such clots sometimes form. Why, 
is not known, "ince inflammatory exudates as a rule do not coagulate. In 
any decomposing alkaline urine, masses which resemble fibrin casts con- 
sisting of pus, mucus and bacteria may be voided or may even plug the 
urinary passages. 

Albuminuria. — Albuminuria or the presence of a coagulable protein 
in the urine, albumin, serum albumin, serum globulin, etc. (see page 
217), may be due to conditions in the renal cortex (true, or cortical 
albuminuria) or to conditions below the cortex (" false " or, better, sub- 
cortical albuminuria). In cases of sub-cortical albuminuria trr^ urine 
as secreted by the kidney cortex is normal and receives the aiDumin 
lower in the urinary passages, either as an inflammatory exudate, lymph, 
blood, or chyle. By albuminuria in the following paragraphs is meant 
that due to some disturbance of the renal epithelium, especially that 
of the glomeruli. 

Albuminuria Without Definite Renal Lesion. — Physiological 
albuminuria, or the constant presence of a proteid in normal urines has 
been mentioned on page 223. Posner first, in 1884, claimed the presence of 
serum albumin in all normal urines, but this was at once doubted, since 
the tests seemed to indicate that the proteid present was mucin or a nucleo- 
albumin. More recent work, however (see page 218), seems to have 
established beyond doubt the presence of a small amount of serum albumin, 
or, according to others of euglobulin and directly from the blood, in prac- 

108 Kolisch and Purion, Zeitschr. f. klin. Med., 1896, Bd. 29, p. 374. 



THE URINE: PROTEIDS PRESENT 225 

tically all normal urines. If we use Speigler's reagent it would be bard 
indeed to find many whose urine is really albumin-free. 

The difference between physiological and pathological albuminurias is 
quantitative not qualitative and the term " albuminuria " now implies 
that serum albumin is present in the urine in such quantities that it can be 
detected by the not very sensitive tests accepted as standard, e.g., the heat 
and acetic acid test is standard in our clinic. Hofmeister's standard was 
that Heller's test should show no ring in 3 minutes. . 

The question of albuminuria is, therefore, similar to that of glycosuria; 
very small amounts of both bodies are found in normal urines but are 
disregarded unless present in amounts sufficient to give the test accepted 
as standard. This line is, however, an artificial one and not very convincing 
to the medical students who are sorely tempted to try the more delicate 
tests on their own urine and are made unhappy as a consequence. 

The problem of physiological albuminuria now little interests the 
clinician. By albuminuria he means albumin in pathological amounts and 
the older discussion of physiological albuminuria must now be continued 
under the title " functional albuminuria." The question now is: May the 
normal person under practically normal conditions pass a urine containing 
enough serum albumin to give the heat and acetic acid test or Heller's 
test within 3 minutes? 

Functional Albuminuria. — The term " functional " albuminuria 
Pavy used merely as a contrast term to " structural " albuminuria in which 
latter case the albuminuria depended on demonstrable anatomical changes 
in the kidney. 

Senator defined an albuminuria as " functional " (he said " physio- 
logical") if it occurs in young men, is transitory, is slight in grade, if the 
further history of that person is negative, if the urine is otherwise normal 
and if its occurrence follows an unusual cause, such as severe muscular 
work by one not used to it ; exposure to cold, nervous stress, or after unusu- 
ally large proteid meals. According to Senator the cause need be an 
unusual one solely for that person and at that time. 

This form of albuminuria was first noticed among soldiers. Leube 
stated that 59% of the raw German recruits showed a temporary albumin- 
uria after a forced march, but not later after they had been strengthened 
by training. Macfarlan 109 found albuminuria in practically every foot-ball 
player for hours after a game, while in the urine of 19 he found granular 
casts as well and in that of 6 also blood-casts and red-blood-cells. Muller n0 
found albuminuria in the urine of 11 bicycle riders after a race and in 
several of these he found also casts of all descriptions and renal epithelium. 
Their urine was normal on the following day. Barach m found in the urine 
of each of 19 Marathon runners albumin, casts, and, in nearly all cases, 

109 ]\j ew York Med. Record, 1894, vol. xvi, p. 769. 

110 Munch, med. Wochenschr., 1896, No. 48. 

111 Arch, of int. Med., Apr., 19 10, vol. v, p. 382. 
15 



226 CLINICAL DIAGNOSIS 

blood. One week later the urine of 4 of these men still contained casts and 
albumin and that of 2 casts alone. In other words, an albuminuria may 
be expected in any athlete no matter how good his condition, if only his 
exertion be strenuous enough and especially if the exertion involves prin- 
cipally the leg and thigh muscles. Normal man in every sphere of life, 
even the trained athletes, have certain limitations and the question arises, 
having overstepped these, can the albuminuria which results be termed 
" functional " even though the persons involved would be considered almost 
perfect physically (trained athletes, et al.)l 

In this same group of functional cases Senator includes the albuminurias 
which follow violent emotions or an unusually heavy proteid meal (ali- 
mentary albuminuria) . Among soldiers, normal men under uniform condi- 
tions, Rapp found that 10.7% showed albuminuria after their mid-day meal. 
Experiments show that the ingestion of excessive amounts of certain pro- 
teids will cause an albuminuria in some apparently normal persons. This 
is especially true if egg albumin be used. Such an albuminuria begins in 
about 2 hours after the meal and lasts about 4 hours. 112 Formerly it was 
claimed that the proteid ingested can itself be detected in the urine but 
the identification of proteins by the specific precipitin method has not 
proven very satisfactory. Some raw egg albumin may be excreted as such 
but even then the most of the protein present will be serum albumin, show- 
ing that the foreign proteid in the blood temporarily at least injured the 
kidneys. Certainly we know that if the kidneys are innammed any indis- 
cretion in the diet may aggravate the condition. An alimentary albumin- 
uria is claimed for the new-born if fed on cow's milk and explained on the 
ground that their intestinal mucosa has not yet developed its normal 
impermeability to foreign proteids. 

Prolonged cold baths will cause an albuminuria. Rem Picci observed 
115 baths taken by 35 healthy men and found that a bath of 3 minutes 
at 12 to 1 3 C, or 15 minutes at 15 to 20 C, caused quite uniformly a 
slight transitory albuminuria, minimal in amounts, never lasting over 
24 hours, together with casts and generally with diuresis with increased 
urea and chloride output. 

Mental over-exertion is said in certain cases to cause albuminuria. 

Albumin is more likely to appear in the urine should several of these 
above-mentioned factors occur simultaneously. The intermittent nature 
of the albuminuria is no proof that it is functional since a truly pathological 
albuminuria may intermit for long periods of time and in a mild case, or 
one during convalescence, the albuminuria may return for but a few hours 
after 1 of the above causes. Certainly the amount is not important 

112 See Ascoli, Munch, med. Wochenschr., 1902, No. 10, and Inouye, Deutsches Arch. 
f. klin. Med., 1902, Bd. 75; and on the opposite side of the question Umber, Berl. klin. 
Wochenschr., July 14, 1902; for the chemical side, Sollman and Brown, Jour, of Exp. 
Med., March 17, 1902. 



THE URINE: PROTEIDS PRESENT 227 

although Senator considers that if it exceeds 0.4 to 0.5 grn. per liter it cannot 
be called functional (he said " physiological"). 

Another example of the so-called " physiological albuminuria " is the 
albuminuria of the new-born. The urine of many infants during the first 
8 or 10 days after birth contains a slight amount of albumin, hyaline casts 
and epithelial cells. Ribbert gave as an explanation that the kidneys at 
birth often are not quite finished; that is, that there is still to occur a 
desquamation of the epithelium of the capsules of the glomeruli. The 
urine found in the bladders of stillborn infants often contains albumin 
and casts, but this may have another explanation. 

The Albuminuria of Women During Pregnancy and in Labor. — 
An alburninuria may be considered as truly physiological which follows 
the confinement. It usually disappears at once. Little, 113 as the result of 
very careful work, concludes that albumin may be demonstrated in the 
catheterized urine of about one-half of all pregnant women, of primiparas 
as often as of multiparae, although casts are found more often in the urine 
of the latter. During uncomplicated labor a still greater number, and espe- 
cially of the primiparse, show albuminuria. This may be due to the 
muscular work and the increased blood-pressure which childbirth involves. 

" Albuminuria of adolescence " (Gull), " of puberty "; " accidental 
alburninuria "; "essential albuminuria"; "physiological albuminuria ; 
" Pavy's disease"; "cyclic albuminuria of the apparently healthy 
" postural," " orthotic," " orthostatic " and " intermittent albuminuria 
From this list of names one may make out the essential features of the 
various cases. Posner proposes the quite satisfactory term, " essential 
albuminuria," for albuminuria is the one and only symptom common to all. 
These cases are of far greater importance clinically than those discussed 
under the title " functional " albuminuria. They form a large group of 
persons who enjoy reasonably good health, but whose urine either con* 
stantly or intermittently contains a trace of albumin. This condition is 
discovered by Army and Navy medical inspectors, by the examiners for 
insurance companies and by the doctors to whom neurasthenics apply for 
treatment. Insurance men say that 5% of all the " normal " persons 
examined while the temperature is above 90 or below o c F. show albumin- 
uria; at other times, only about 2%. 

Of the group as a whole it may be said that these individuals are for 
the most part youths or young adults who are not robust. Some are anemic 
and many complain of symptoms referable to instability of the vasomotor 
system. Many come of neurotic families while more have had those infec- 
tious diseases which often are followed by nephritis; such as tonsillitis, 
scarlet fever, diphtheria, etc., although they have no signs or symptoms 
of kidney trouble other than an albuminuria which may be continuous, 
or if intermittent appears in response to some ordinary acts of eve^day 

1,3 Amer. Jour, of Obstet., vol. i, No. 3, 1904. 



228 CLINICAL DIAGNOSIS 

life ana not to unusual causes. The tendency now is to eliminate from this 
group all in whom latent infection can be demonstrated, whether of the 
tonsils, teeth, nasal sinuses, gall-bladder, appendix, tubes or prostate (some 
would add the bronchial tree and the colon wall) on the ground that these 
patients have an actual subacute nephritis; also, all those who are anemic 
(and so probably harbor a latent infection) or have had recently an infec- 
tious disease. Essential albuminuria is sometimes a family disease present 
in 2 or even 3 children of one family. In this group some include the albu- 
minuria of masturbators and that which follows sexual excitement. In 
these cases the albumin often is present only in the morning specimen. 
Some (Sir Andrew Clark and others) say that this proteid is from a secre- 
tion of the urethra or accessory sexual glands. 

A diagnosis of essential, etc., albuminurias is possible only after a long, 
careful study of the individual case, including his past history, a careful 
physical examination especially of the circulation and eyes, an accurate 
study of the urine with especial reference to its specific gravity, amount, 
sediment, etc., all have failed to reveal any other evidence of renal disease 
and even then an autopsy may reveal a true nephritis. The fact that the 
albuminuria is intermittent or is orthostatic, or is, or is not, accom- 
plished by a cylindruria, does not help to exclude a latent acute nephritis 
during convalescence. 

Leube taught that the cases of albuminurias of adolescence formed a 
separate, distinct group. In them the albuminuria is present only between 
the ages of 14 and 18 years and then disappears. It is attributed to an 
insufficiency of the kidneys relative to the growing organism, associated 
with instability of the vasomotor centers. In this group are found most, 
but not all, of the cyclic or postural cases. In them the element of heredity 
would seem to be important. The cases reported by Lommel 114 would fall 
under this title. He reported that of 587 factory workers from 14 to 18 
years old, 18.9% showed slight albuminuria once or many times and either 
no sediment, or at the most a few hyaline casts and fatty epithelial cells 
in the centrifugalized specimen; of 130 patients from the same class but 
over 25 years old, only 1 showed albuminuria. Functional cardiac and 
vascular disturbances were common among these workers. Posner empha- 
sized sexual excesses at puberty as a common cause of this form of albumin- 
uria. Sutherland 115 noted a definite relationship between albuminuria 
and the movable kidneys present in % of his cases. 

Cyclic albuminuria is the most interesting of all so-called physio- 
logical albuminurias. In these cases one can demonstrate a remarkable 
daily cycle; the albumin is absent at night and when the patient lies flat 
on his back but appears whenever he stands up. The terms " orthostatic " 
or " postural " albuminuria therefore are much preferred by some for this 

114 Deutsches Arch. f. klin. Med., 1903, Bd. 78, p. 541. 

115 Am. Jour, of the Med. Sci., 1903, vol. cxxvi. 



THE URINE: PROTEIDS PRESENT 229 

group although by usage the term cyclic albuminuria implies one not due 
to nephritis, while the best examples of orthostatic albuminuria are seen 
in cases of mild nephritis. 

The cyclic cases may be subdivided into 3 groups: those associated 
with vasomotor phenomena (the neuropathic element predominating), 
those with abnormal renal circulation, as in congenital floating kidney, 
and the hereditary form. Mix also 116 recognized an intermittent and a 
continuous type of cyclic albuminuria. In these intermittent cases the 
albumin as a rule appears after rising, reaches a maximum at noon or from 
4 to 6 p.m., then declines and disappears from 8 to 10 p.m. If the patient 
changes his habits the cycle will change also. In many cases the albumin- 
uria bears little or no relation to meals. The cycle affects not only the 
albuminuria but also the water output (which decreases as the albumin 
appears). As a rule one can note the following sequence : first, an increase 
in the pigments, then the appearance of albumin, then an increased output 
of uric acid and lastly an increase of urea (Teissier). While casts are rare, 
yet careful search will, as a rule, discover a few. This albuminuria may 
even be diminished by exercise and fatigue. The cycle in the continuous 
form continues for years and if it ceases does not return. These cases 
practically never develop into Bright 's disease. The adults of this group 
are neurasthenics with vasomotor instability and 37.5% of the children 
have a congenital movable kidney. Armstrong 117 found this form of 
albuminuria in 12 to 15% of over 3000 school boys. It is seen more in 
summer than in winter; heredity is often present; it is often associated 
with depression of spirits and fainting spells, especially while the boy is 
idle, not when occupied; the boy is apathetic, his heart is subject to inter- 
mittent attacks of dilatation and palpitation; it lasts only during puberty. 

It is an open question whether cyclic albuminuria should be considered 
as physiological or pathological. When this discussion began nephritis, 
no matter how slight, was thought of as a disease which, once begun, was 
likely to continue for years or even for life. With better knowledge of 
the importance of latent infections in the production of nephritis and the 
excellent prognosis when these cases are properly treated, one reason for 
the use of the term " cyclic albuminuria," has in large measure disappeared. 
Most transitory albuminurias certainly are pathological, e.g., those of 
fevers. Posner's case was well after 17 years. Senator 118 insisted that the 
majority of these cases have no nephritis. Krehl, who followed several 
cases over a long period of time, considered this condition harmless and 
not a form of mild nephritis. Broadbent 119 has never known a true case 
of this form to develop actual rental disease. In all the above cases the 
amount of albumin is small, the amount of urine normal and its specific 

116 Am. Jour, of Med. Sc, 1904, vol. cxxviii, p. 307. 

117 Brit. Med. Jour., 1904. 

118 Deut. Arch. f. klin. Med., December 8, 1904. 

119 Brit. Med. Jour., 1904. 



230 CLINICAL DIAGNOSIS 

gravity normal. A few hyaline casts may or may not be present. There 
are no cardiovascular changes. The immediate cause is much in dispute. 
Possibly the most reasonable explanation will be one which associates the 
urinary findings with changes in the renal circulation which follow changes 
in posture. Edel found in 3 very interesting cases that the albumin-free 
intervals (in the afternoon as a rule) were also periods of diuresis and that 
in some degree the amount of albumin varied inversely to that of the urine. 
Erlanger and Hooker 120 found that the amount of albumin varied inversely 
as the pulse-pressure. Many of these cases later recover. The last and 
best review of this subject is that of Hooker. 121 

The diagnosis of cyclic albuminuria often can be made only after years 
of observation, since many patients who first are included in this group 
later prove to be cases of true Blight's disease. 

The hypostatic albuminuria of splenic origin seen in some patients 
with enlarged spleens while recumbent and absent while they are erect, 
is, Rolleston thinks, the opposite of cyclic albuminuria. Since it is not seen, 
however, in all patients with enlarged spleens nor in those whose spleens 
are largest, some other causal factor is necessary. Pressure on the left 
renal vein may explain some of the cases. This may resemble the albumin- 
uria in chronic passive congestion of mitral disease. 

Albuminuria Minima (Lecorche and Talamon). — Under the heading 
" albuminuria minima " are included cases whose urine constantly contains 
a trace of albumin, but almost never over 0.5 gm. per liter. The output 
varies very little with the position of the patient, the time of day, diet, etc., 
although for each patient there may be some individual factor which in- 
creases the output. Some of these cases are quite certainly convalescent 
from a latent subacute nephritis. The prognosis is uncertain and must be 
guarded, for some later are cases of a true nephritis. Others remain the 
same for years with no further symptoms. Under this group the French 
put the post-infectious cases, albuminuric residuale, albuminuric paracel- 
laires (or insular nephritis), albuminuric cicatricielle (due to imperfect 
healing, leaving a " scar ") ; also the albuminuria of adolescence, the heredi- 
tary form, albuminuric phosphaturique, and the albuminuric pregoutteuse. 

Intermittent albuminurias are those which persist for periods lasting 
weeks, months or years and then cease for longer or shorter periods. This 
term does not include the cyclic or postural albuminurias in which the 
albumin-free periods last for hours only. They usually are cases of insidious 
nephritis which have followed some preceding acute infectious disease. 
In other cases the albuminuria, with casts often, is due to and is present 
only during periods of broken cardiac compensation. 

The intermittent heredity form of albuminuria includes, according 
to some, many patients who formerly showed the albuminuria of adoles- 

120 Johns Hopkins Hosp. Rep., vol. xii, 1904. 

121 Arch, of Inter. Med., 1910, vol. v, p. 491 



THE URINE: PROTEIDS PRESENT 231 

cence, but who now in adult life are albumin-free except in response to 
fairly adequate causes. 122 In some cases the parents of the patients had 
had albuminuria during youth, while in others a neurotic family history is 
the only suggestive feature. 

Traumatic Albuminuria. — Transitory albuminuria may follow injury 
to the brain, as apoplexy; injuries which crush the kidneys, in which cases 
the albuminuria and cylindruria may continue for a long time without the 
appearance of other signs of nephritis, although this may explain some 
cases of so-called benign latent contracted kidneys (Stern, Curschmann) ; 
bimanual palpation of the kidney, even that of an ordinary physical 
examination (Menge 123 could in 15 of 21 cases cause by bimanual palpa- 
tion a transitory albuminuria lasting usually from 1 to 24 hours and in 
some cases also a slight hematuria) ; and finally anything obstructing renal 
venous flow, as in movable kidney during Dietl's crises. 

Febrile Albuminuria. — During any acute fever, but especially pneu- 
monia, typhoid, malaria, acute articular rheumatism, grippe and acute 
tonsillitis, there may be a slight albuminuria which begins with the fever 
and disappears with it. The renal lesion in such cases is thought to be the 
cloudy renal epithelium, the faintest grade of inflammation (Leyden). 
The amount of albumin in the urine of these patients usually is small, 
but sometimes is great. Hyaline and epithelial casts are often found but 
no other formed elements suggesting inflammation. These cases differ 
from true cases of nephritis only in degree. 

Under the term hematogenous albuminuria is included a very large 
and confusing group of non-febrile cases which show at autopsy no renal 
lesions, except, perhaps, slight parenchymatous changes. In this group 
occur some of the cases of purpura, scurvy, chronic lead or mercury poison- 
ing, lues, leukemia, cachexia, severe anemia, cholemia, hyperglycemia and 
ether and chloroform narcosis. 

" Hematogenous albuminuria," accurately speaking, would indicate 
one due to the elimination by normal kidneys of some proteid in the blood 
which cannot be used. Cases in which there is a possibility that the renal 
epithelium has been injured, as by the proteid itself or some poison as lead, 
mercury, etc., should be excluded. It is true that foreign proteids, e.g., 
albumoses, egg albumin, peptone, casein, free hemoglobin, etc., if intro- 
duced into the serum will appear in the urine and formerly this was the 
explanation (an unfit proteid) of all forms of albuminuria, but the chances 
are that all are due in part at least to some disturbance of the renal cells 
themselves, for they certainly are exceedingly sensitive to changes in their 
nutrition. Indeed a true hematogenous albuminuria has not yet been proven. 

The Nervous Form. — Patients with epilepsy, apoplexy, tetanus, 
exophthalmic goiter, injuries to the head, delirium tremens, various psy- 

122 Dieulafoy, Loude, Arch. gen. de med., n.s., ii, 3, p. 257, 1899. 

123 Munch, med. Wochenschr., June 5, 1900. 



232 • CLINICAL DIAGNOSIS 

choses, even neurasthenia and migraine, often have a slight transitory 
albuminuria. In some cases a few casts also appear. In other very inter- 
esting cases casts and no albumin are found. We followed for several 
weeks the urine of such a case, a boy 14 years old with hysterical attacks 
followed by cylindruria. A very transitory early albuminuria could not be 
excluded, but not one of the specimens examined contained albumin. 

Closure of the ureter, retention of urine in the bladder, compression of 
the thorax, have been accompanied by albuminuria; digestive disturbances, 
as obstruction of the bowel (a reflex cause being assumed as in cases of 
strangulation of the bowel or omentum 124 ), acute diarrhea, constipation 
and liver disease are sometimes given as causes. In % of the cases of 
intestinal obstruction with albuminuria the albumin disappeared after the 
obstruction was relieved even though the bowel had become gangrenous. 
The cause in these cases of the albuminuria is uncertain but it certainly 
was not any attending peritonitis. 

Albuminuria with Definite Renal Lesions. — An active renal congestion 
such as that due to exposure to cold, or in children to the acute infections, 
and chronic passive congestion due to heart or lung disease, to tumors, or 
to pregnancy, usually produce an albuminuria. The albumin in such 
cases is small in amount and runs parallel to the amount of urine, while 
in cases of true nephritis its percentage varies inversely as the amount 
of urine. 

Nephritis. — All cases of nephritis at some time during their course 
produce an albuminuria and in general the more acute the nephritis the 
larger the percentage of albumin, but not the larger the total amount of 
albumin per day. For, to excrete a larger amount of urine with a lower 
percentage of albumin is evidence of a better renal condition than when 
the percentage is higher but the total output of urine and so of albumin is 
smaller. It is not true that the prognosis of the case is always determined 
by the acuteness of the nephritis. In very young persons this may be the 
case but in nearly all adults with so-called acute nephritis the kidneys 
suffer also from previous attacks of this disease, or from years' long con- 
tinued attacks, which have led to extensive destruction of renal epithelium. 
In the renal disturbance present at anyone time, the acute element, of which 
the albuminuria is a good index, may be relatively little. It is for this 
reason that in cases of chronic interstitial nephritis ending in uremia and 
death the urine may contain but mere traces of albumin. In severe cases 
periods of albuminuria may alternate with months during which the urine 
is quite clear of demonstrable protein. In some cases of definite acute 
nephritis there may be no albuminuria. 125 In nephritis the urine seldom 
contains more than 1% of albumin. Sometimes it contains 2%, in very 
rare cases 5%, and in 1 case 8%. Senator mentions a case of subacute 

124 Neumann, Trans. Clin. Soc. of London, 1897, Bd. 30, p. 65. 

125 Herringham, Trans. Clin. Soc. of London, vol. xxxiv, p. 901. 



THE URINE: PROTEIDS PRESENT 233 

nephritis whose urine, for a period covering several days, contained from 
6 to 8% of albumin. These cases with large amounts of albumin are, 
interestingly, often luetic. There are between 20 and 25 such cases of 
nephritis syphilitica acuta precox on record. In Hoffmann's case the 
enormous albuminuria ran parallel to the luetic symptoms and improved 
under mercurial treatment. In amyloid disease of the kidney the amount 
of albumin may be great or veiy small . As a rule it varies from o . 5 to o . o 5 % . 

Salkowski reported a case l26 the urine of which had a specific gravity of 
1.056 and contained 7% of proteid. On standing, a rich, white, amorphous 
precipitate, not a coaguhim, was deposited which gave the chemical reac- 
tions of albumin. On another day the urine of this patient contained even 
8.5% of albumin. (The blood contains only about 7.5% of proteid.) 

The total daily output of albumin in cases of nephritis is usually small, 
from 1 to 20 gms. Nephritis is not serious because of the actual loss of 
albumin to the body, for this could as a rule easily be covered by a very 
slight increase in the diet. 

The albuminuria of a case of nephritis can be judged accurately only 
if the patient is on a carefully controlled diet and if the curve of the daily 
albumin, water, and nitrogen output are followed for at least 3 weeks. The 
customary methods of urine examination and the clinical uses made of the 
results of these examinations show an amazing lack of intellectual honesty 
or of downright ignorance. These curves show marked waves due appar- 
ently to changes in the temperature of the room, its humidity, the baro- 
metric pressure, others due to the mental condition of the patients, others 
due to changes in the diet, etc., but all too complex for off-hand analysis. 
In some cases thus studied it would seem as though meat were not as harm- 
ful as vegetables, some cases seemed to improve if salts are prohibited, 
some if used. In most cases the albumin is decreased (other things practi- 
cally equal) if the patient is on a milk diet abundant enough to cover tissue 
needs. In practically all cases the albumin is increased by the erect posture, 
for all cases of albuminuria are relatively orthostatic, but this does not 
explain its increase during the waking hours of those patients who because 
of hydrothorax or cardiac disease are propped up against a back rest at 
about the same angle day and night. Exercise of any kind, even massage 
(Edgren) , increases the output. As a case improves the value of an increase 
in diet and an increase of exercise will be proven by a diminution of 
the albuminuria. 

" Hetero-albumosuria ; " Bence-Jones Body; "Kahler's Disease;" 
" Myelopathic Albuminosuria of Bradshaw ; " Theromolytic Albuminuria. — 
A remarkable proteid formerly called "albumose" appears in the urine of 
certain patients in very large amounts. It was supposed because of some 
of its chemical properties to be an hetero-albumose. Magnus-Levy 
however, who obtained it in crystalline form, showed that among its 

126 Berl. klin. Wochenschr., March 3, 1902. 



234 CLINICAL DIAGNOSIS 

digestion products are all the primary proteoses except hetero-albumose. 
It would seem to belong in a group by itself standing nearest the true 
albumins. The safest course is, therefore, to call it " Bence-Jonesbody." 
In 1903 but about 35 well studied cases had been reported, all but one of 
which (a patient with lymphatic leukemia (Askanasy)) were cases of 
multiple myelomata. Later Boggs and Guthrie 127 report the condition in a 
case of carcinoma with metastases to the bone-marrow. In all the cases 
reported there was extensive disease of the bone-marrow. The most of 
the patients died in less than 2 years from the discovery of the condition. 

The Bence- Jones body is present in the urine often in large amounts, 
even 7%, but in the majority of cases there is less than 1%. Some cases 
are reported as intermittent (Boston). Coriat 128 reported a case with 
none in the urine but with 4% in the pleural fluid. 

Reactions. — If urine containing the Bence- Jones body is first acidified 
with acetic acid and then warmed there will develop at a low temperature 
(about 6o° C., often 52 ) a milky, then a heavy, sticky precipitate which 
will for the most part, and in some cases perfectly, disappear on bringing 
the urine to a boil, but which will reappear on cooling. This is the charac- 
teristic reaction, and suggested the name " thermolytic albuminuria " 
proposed by Hugounenq for this condition. 

Another very important test is the following. If nitric acid be added 
to the urine a heavy precipitate forms, which is soluble on warming and 
reappears at cooling. The urine will give the biuret reaction. 

These striking reactions should attract attention at once. The urine 
may be turbid when voided. The moderately low temperature at which 
the precipitate appears, in general below 6o° C., depends on the amount 
of Bence- J ones body present and also on the salt content of the urine. 
The properties of the Bence- Jones body found have been described so 
differently that it might seem as though we were dealing not with one but 
with a group of bodies. The chances are, however, that these differences 
are due to the varying amounts of salts and urea in the urine. 

Boston 129 proposed the following test for the Bence-Jones body, based 
on the large amount of loosely bound sulphur it contains. From 15 to 
20 c.c. of urine are mixed in a test-tube with an equal amount of saturated 
NaCl and well shaken. Then 2 to 3 c.c. of 30% NaOH are added and the 
tube shaken hard. The urine at the top of the tube is then heated to boiling 
and PbAc solution (ic%) added drop by drop, heating the urine after the 
addition of each drop. In % to 1 minute there will develop a brown, later 
a black, precipitate. 

The daily amount of this body eliminated is quite constant and since 
it is not affected by diet it probably is not a non-assimilable product of 

m Bull, of the Johns Hopkins Hosp., Dec, 19 12. 

128 Am. Jour. Med. Sci., 1903, vol. cxxvi. 

129 Am. Jour. Med. Sci., 1902, vol. cxxiv. 



THE URINE: PROTEIDS PRESENT 235 

digestion but rather a substance formed in the bone-marrow, some say 
from the granules of the myelocytes and tumor-cells. 

This albumose can be demonstrated also in the ascitic fluid, blood and 
bone-marrow of patients with this disease. 

Quantitatively the Esbach's tube method will give an approximate 
determination (see page 216). 

Albumosuria, " Peptonuria." — Under the term albumcse have been 
described at least 2 different groups of bodies, " Bence-Jones body," and 
several digestion products formerly called " peptones " (a name given by 
Briicke to proteids not precipitated by K 4 FeCN 6 and acetic acid) but now 
identified as a mixture of the deutero-albumoses and the peptone of Kuhne. 
(Kuhne defined a peptone as a proteid not precipitated by complete satura- 
tion with (NH 4 ) 2 S0 4 and yet which gives the biuret test.) The true pep- 
tone (of Kuhne) always with albumose (the reverse is not true), has been 
identified in the albumosurias of croupous pneumonia, of ulcer of the stom- 
ach, of pulmonary tuberculosis and of the puerperium. 130 

In testing for the deutero-albumoses the urine should be made albumin- 
free by the Hofmeister method, Morner's body removed by basic lead 
acetate and the urine then saturated with ammonium sulphate. A floccu- 
lent precipitate indicates albumose (Hofmeister). A better method is to 
add to the urine (cleared of albumin and of Morner's body) % volume of 
concentrated acetic acid and then phosphotungstic acid. The precipitate 
is dissolved in a little water, NaOH or KOH are added in excess and then 
very dilute CuS0 4 . It may be necessary to filter off the precipitate of 
Cu(OH) 2 . Deutero-albumoses are indicated by the appearance of a violet - 
red color. 

Hammarsten recommends a method which, as modified by Bang, is of 
clinical value. Ten parts of urine plus 8 parts of saturated ammonium 
sulphate are boiled for a few seconds and this hot fluid then centrif ugalized 
from y 2 to 1 minute and decanted. The urobilin is then extracted from the 
precipitate with alcohol, the residue then taken up with little water, heated 
to boiling and filtered. In this way the albumin is removed. The filtrate 
is then shaken out with chloroform to remove the last trace of urobilin, 
the chloroform pipeted off and the biuret test applied to the remaining 
aqueous solution. 

Alder 131 recommended the following test as more accurate. Albumin 
if present is removed by trichloracetic acid (15%). To from 6 to 10 c.c. 
of urine in a test-tube are added 1 or 2 drops of HC1 till acid, then 5% 
phosphotungstic acid till precipitation is complete. The fluid is then centri- 
fugalized for a few seconds. The supernatant fluid is poured off, the sedi- 
ment suspended in absolute alcohol and again centrif ugalized. This is 
repeated till the sediment and the alcohol (colored yellow with urobilin) 

130 Ito, Deutsches Arch. f. klin. Med., 1901, vol. lxxi. 

131 Berl. klin. Wochenschr., 1899, pp. 764, 780. 



236 CLINICAL DIAGNOSIS 

are white and clear. The sediment is then suspended in water, strong 
NaOH added, the fluid shaken till all blue color disappears, then CuS0 4 
solution is added. By this method even 0.2 gm. of albumose per liter can 
be detected. 

The deutero-albumoses may appear in the urine alone or with albumin. 
Albumosuria is as a rule clinically important only if the urine is quite free 
of albumin. In nephritis, albumose is often present with the albumin, but 
it may precede it or continue after the albumin has disappeared. Its 
presence in nephritis formerly was interpreted as due to the products of 
digestion which escape through the kidneys but now is attributed to a 
pepsin -like ferment which is often present in the urine. The albumo- 
suria often present in luetic nephritis may arise in gummata under- 
going involution. 

Hematogenous Albumosuria. — When there is considerable albumose 
in the blood some at least will be excreted in the urine, but not if the amount 
in the blood is small. Albumosuria may therefore accompany any condi- 
tion with disintegration of a tissue or exudate, or any disease with increased 
catabolism, as cancer or the fevers. It occurs therefore in leukemia, scurvy, 
purpura, in cases poisoned by a hemolytic poison or by a toxin which 
destroys tissue-cells. The albumosuria of the puerperium is ascribed to 
the involution of the uterus; that following the death of a fetus to the 
maceration of the infant; but albumosuria occurs also in some cases of 
normal pregnancy. 

Enterogenous albumosuria develops in some cases of gastric or 
intestinal ulcer, as, e.g., in intestinal tuberculosis. In such cases the inges- 
tion of small amounts of certain albumoses, e.g., of somatose, will be fol- 
lowed by an albumosuria, but usually large amounts must be administered 
(alimentary albumosuria). The attempt to make use of this as a test of 
ulcer has not been very successful. (The patient was fed from 40 to 60 gms. 
of albumose. If an albumosuria followed the presence of a gastric or 
intestinal ulcer was assumed.) 

" Hepatogenous albumosuria " develops in acute yellow atrophy, 
in cirrhosis of the liver and cancer of the liver, in catarrhal jaundice and 
in phosphorus poisoning. " Febrile albumosuria " is met with in the 
infectious fevers as the temperature falls, in rheumatism, septicemia, 
typhoid, phthisis, gangrenous processes, measles, scarlet fever, erysipelas 
and smallpox. It occurs in some mental diseases and in paralytic 
conditions. " Pyogenic albumosuria " is supposed to accompany the 
absorption of an exudate, as in pneumonia during resolution, in em- 
pyema, bronchiectasis, epidemic cerebrospinal meningitis, abscess and 
in osteomyelitis. It may be met with in gangrenous processes or cancers 
of any organ. 

One should of course exclude the albumose of spermin and that of the 
secretions of the accessory genital glands. 



THE URINE: PROTEIDS PRESENT 237 

Since albumosuria is met with in so many conditions it can have rela- 
tively little clinical value, yet it may be of some service in the recognition 
of a suspected abscess (e.g., of the appendix, brain, or an empyema) and 
in the differential diagnosis between tuberculosis and epidemic cerebro- 
spinal meningitis. 

Hematuria. — Hematuria may be met with under the following conditions : 
(i) General diseases: the malignant forms of acute specific fevers, 
especially smallpox, typhoid fever and malaria; in leukemia occasionally; 
in the so-called hemorrhagic diathesis, as hemophilia, scurvy, morbus 
maculosus Werlhofii and the purpuras. In the latter diseases the process 
may even be limited to the kidney. 

(2) Renal causes : acute and chronic passive congestions and inflamma- 
tions of the kidney; all forms of nephritis at the onset and practically 
every form of nephritis at some time during its course ; nephropathies due 
to turpentine, carbolic acid and cantharides, especially at the onset; sub- 
acute parenchymatous nephritis always, Weigert said; in renal infarctions, 
although marked hemorrhages are rare; in new growths of the kidney, in 
which cases the hematuria sometimes is profuse; at the onset of renal 
tuberculosis, especially if the papilla? are involved; in cystic kidneys, 
renal calculus and, lastly, in parasitic diseases of the kidney, especially 
that due to filaria, echinococcus and Distoma hematobium. In con- 
gestion due to venous thrombosis, e.g., of the new-born, hematuria is said 
to be common. 

(3) It may be due also to lesions or diseases of the urinary passages: 
e.g., stone in the pelvis or ureter, tumors and ulcers of the bladder, parasites 
of the bladder, calculi, ruptured veins and urethritis. 

(4) In trauma of any part of the urinary tract from the kidney down. 

(5) And, lastly, there is an interesting group with no known lesion, the 
so-called " Gull's renal epistaxis," " essential renal hematuria," " angio- 
neurotic hematuria," or " renal hemophilia." This is a rare disease of 
middle adult life. The hemorrhage in these cases may be unilateral. In 
a few of these cases angiomata of the kidney have been found, in others 
no' gross lesions and so nervous causes are suspected. Some of these cases 
recover without any special treatment, others after treatment for a neu- 
rosis, while others after a nephrotomy, a nephropexy or the simple exposure 
of the kidney. 132 Eshner 133 collected 48 cases of unilateral renal hematuria, 
most of which had been diagnosed as calculus or cancer. Since then other 
interesting cases have been reported. 

Recent more careful examinations have thrown considerable doubt on 
the normal condition of these kidneys and only too often has the diagnosis 
of " essential hematuria " been the confession of ignorance and done great 
harm. A diagnosis of unilateral hemorrhagic nephritis was made in Stich's 

132 Stavely, Johns Hopkins Hosp. Bull., March, 1893. 

133 Am. Jour. Med. Sci., 1903, vol. cxxv. 



238 CLINICAL DIAGNOSIS 

case. 134 In Schuller's 135 case the kidney looked normal on gross examination, 
but microscopically a chronic parenchymatous nephritis was demonstrated. 

In conclusion it may be said that the most frequent causes of profuse 
urinary hemorrhage are stone, tumor and tuberculosis, and in cases of 
profuse painless hemorrhage tumor especially must be excluded. 136 

It is customary to limit the term " hematuria " to those conditions 
with the blood grossly visible in the urine. Such urine is always turbid 
and has a color which varies from a light smoky tint to a bright red or 
blackish-brown. Microscopically the red blood-cells are found in various 
states of preservation, as well as other elements which will vary according 
to the cause. In renal hematuria the blood seldom clots, but is homo- 
geneously mixed with the urine while in cases of vesical hemorrhage the 
second glass of the 2 -glass test will contain more blood than the first. If in 
a case of bleeding from the bladder this organ be irrigated, all the washings 
will be blood-stained, while in renal cases some will be clear. In the acute 
exacerbations of a subacute parenchymatous nephritis especially the 
amount of blood in the urine may be considerable. Blood clots are passed 
in cases of trauma, aneurism, or varices, and in cancer more often than in 
calculus. Sometimes the shape of the clot will betray its origin in the renal 
pelvis or the ureter. 

Gerhardt thinks that if the hemorrhage is from the renal cortex the 
red cells in the urine are more apt to be fragmented or more spherical and 
more leathery in color than usual, and will be accompanied by casts of 
various kinds, especially blood-casts, or casts with red cells attached, and 
renal epithelium. 

In women the vagina must always be excluded as the source of the 
blood. 

Hemoglobinuria or the presence of free hemoglobin in the urine will 
develop whenever the destruction of red blood-cells is so great that the body 
cannot warehouse the liberated hemoglobin. This will be the case when 
about %o or more of the total hemoglobin is set free at one time. Such a 
condition may be due to various blood poisons, as potassium chlorate, 
pyrogallic acid, CO, naphthol, AsH 3 , etc. ; or to the poisons of fevers, especi- 
ally malaria and lues, but also scarlet fever, typhoid, yellow fevers, etc.; 
or to severe burns, exposure to cold, or the transfusion of an incompatible 
serum. It may occur during pregnancy (Brauer), as an epidemic fever of 
the new-born, in certain cases of nephritis and after severe intra-abdominal 
hemorrhages. 

The best known form of hemoglobinuria is the black-water fever seen 
in malarial countries. Curry, Brem 137 and others have shown that while 

134 Mitth. aus. d. Grenzgeb. d. Med. et Chir., 1904, Bd. 13, p. 781. 

135 Wien. klin. Wochenschr., 1904, No. 17. 

136 Kretschmer, The Joui . of A. M. A., Feb. 24, 1917, lxviii, p. 578. 

137 Jour. Am. Med. Assoc., May 3, 1902; Brem, Jour, of A. M. A., Dec. 8, 1906; Love- 
lace, Arch, of Int. Med., June 15, 1913, vol. xi, p. 674. 



THE URINE: PROTEIDS PRESENT 239 

many cases of this condition are quite certainly due to malaria, yet it is 
not always possible to prove this by demonstrating malarial parasites 
either in the blood or the internal organs. Black-water fever may recur 
later after an average dose of quinine. 138 In this disease the hemoglobinuria 
is always accompanied by an intense albuminuria. These may appear 
synchronously, but the albuminuria may persist after the hemoglobinuria 
has disappeared (Brem) . It is the general belief that a hemoglobinuria is 
always the result of a hemoglobinemia, although this has never been proved 
in the hemoglobinuria of infectious diseases or of hemorrhagic nephritis. 

Paroxysmal hemoglobinuria is an interesting and rare disease of adults 
the chief symptom of which is an hemoglobinuria following exposure to 
cold or exertion, often preceded by fever, with chills and pain in the 
lumbar regions, which may last for a few hours or even for 2 days. This 
hemoglobinuria is usually preceded by a hemoglobinemia. 

The urine in this condition, which should be examined while very fresh, 
may be clear but it is usually more or less clouded by hemoglobin casts, 
amorphous masses of pigment and by casts due to the associated nephritis. 
When centrifugalized, the supernatant urine is a clear, blood-colored fluid, 
while the sediment contains very few red blood-cells but amorphous blood- 
pigment in masses or casts, and even crystals of hematoidin. Often hyaline 
and granular casts and renal epithelium are present and sometimes many 
calcium oxalate crystals. The urine of a case of hematuria not examined 
fresh may contain much hemoglobin in solution since the red blood-cells 
will quickly lake but the sediment of such a specimen will be abundant, 
grayish-brown in color and in it may be seen the stromata of the many 
laked red cells. 

With the spectroscope one will find in the urine of a case of paroxysmal 
hemoglobinuria met hemoglobin alone or this together with hemoglobin. 
Serum albumin is always present in the urine and often bile pigment, but, 
it is said, never any bile acid. After the hemoglobin disappears the albu- 
minuria may continue for a short time. Other features of the blood during 
an attack are a leucocytosis and a great increase of the number of platelets. 
Sometimes the hemoglobinemia is evident from inspection of the plasma 
of a specimen of centrifugalized blood. 

Among the common immediate causes of the attacks of paroxysmal 
hemoglobinuria are excessive exercise and mental excitement but exposure 
to cold is the most important. Some patients can induce an attack by 
plunging the hand into cold water and Homburg's patient 139 showed it 
after an involuntary cold plunge of 3 minutes duration. It mRy be pro- 
duced locally by tying a string about 1 finger. Some claim that the cause 
of these attacks of hemoglobinemia is the hemolytic action of the patient's 
own blood-plasma; 140 others, a chemical toxin; others, an increa sed sus- 

138 Nansen, Brit. Med. Jour., May 16, 1903. 

139 Zeitsch. f. klin. Med., vol. liii. 

140 Hoover and Stone, Arch, of Int. Med., Nov., 1908, vol. ii, p. 392. 



240 CLINICAL DIAGNOSIS 

ceptibility of the cells to mechanical injury (and in the circulation shadows 
are found) ; others, including Senator, think the cause is in the kidney (see 
page 328) ; others say lues and it is of interest that at least 23 of the 77 cases 
reviewed gave a history of lues : but the best explanation is that of Moss 
(see page 529). 

Tests for Hemoglobin in the Urine. — Whether or not hemoglobin 
is still within its cells the microscope alone can decide. To determine the 
chemical form of hemoglobin and its modifications, spectroscopic examina- 
tion is necessary. The chemical tests are the same for hemoglobin and 
all of its modifications. 

(1) The presence of blood in the urine may be suspected if the coagulum 
formed by the ordinary heat-acetic-acid test for albumin is brown, swims 
on the surface and is decolorized when shaken out with alcohol acidified 
with H2SO4. This test is not very delicate. 

(2) Heller's Test. — The urine, which half fills a test-tube, is made 
strongfy alkaline with about 5 drops of NaOH and then warmed in order 
to transform all hemoglobin present to hematin. The precipitate, consist- 
ing of the phosphates and carbonates of the alkaline earths, which form 
under these conditions will carry down all the hematin and therefore have 
a brownish red or blood-red color. If the urine originally used was already 
alkaline the phosphates of the alkaline earths may already have precipi- 
tated, in which case it is necessary to add a certain amount of normal urine 
in order to supply these salts. 

This test is very delicate, indicating as it does 1 c.c. of blood in 1 liter 
of urine. If the fine red color of the precipitate is masked by the dark color 
of the urine or by bile the precipitate should be filtered off and dissolved 
in acetic acid. A red solution is obtained which will decolorize gradually 
in the air. This red precipitate has by reflected light a greenish tinge. If 
but little hematin be present the precipitate should be freed of all inorganic 
salts by dissolving them in acetic acid and the residue used for Teichmann's 
test. Precipitates similar in color may be obtained after the ingestion of 
senna, rhubard, or rhamnus. The urine in these cases, however, should be 
yellow at first and red on the addition of sodium hydrate. Hematoporphy- 
rin and other pathological pigments may give a red precipitate, but the 
spectroscope will quickly indicate the difference, for if blood was present 
we now would get the spectrum of alkaline hematin. 

If but a trace of blood is present the urine may first be made alkaline 
with NH4OH and then precipitated with tannic acid. The precipitate 
obtained is used for the hemin crystal test. 

(3) Teichmann's HCl-Hemin Test. — The precipitate obtained by either 
of the preceding methods, or, better still, a tannic acid precipitate, is 
filtered, washed and thoroughly dried in the air. A very small granule of 
this dry precipitate is put on a slide with 1 minute granule of NaCl and a 
few drops of glacial acetic acid and covered with a cover-glass. The speci- 



THE URINE: PROTEIDS PRESENT 241 

men is then warmed over a small flame so that the acetic acid will just 
steam and not boil, renewing the acid as rapidly as is necessary. When 
the hot acetic acid surrounding the granule has taken on a brownish color, 
the heating is discontinued and the slide allowed to cool slowly. The 
characteristic crystals of hemin may soon be seen. Excellent specimens 
may be obtained without the use of heat if the specimen is allowed to stand 
for 24 hours (see Fig. 36). 

This is a very good test to use as a class exercise since it trains the 
student in a valuable and much neglected technic. 

(4) The tannic acid precipitate mentioned in test (2) may be ashed on 
a platinum foil, the ash dissolved in a few drops of hot HC1, this diluted and 
filtered and tested for iron with the potassium ferrocyanide solution. 

(5) The Guaiac test (Schonbein-Almen test) is very delicate. The urine 
acidified with acetic acid if necessary, is overlaid carefully with a mixture 
of equal parts of guaiac tincture (alcoholic solution of resina guaiaci, 1 to 5) 
and old oxygenated oil of turpentine. The tur- 
pentine should previously have been exposed to 

the air for some time that it may be well oxygen- 
ated; the guaiac tincture should be kept in a 
colored bottle and not unnecessarily exposed to 
the sun or air. These solutions when mixed should 
not develop any trace of a blue color. If blood 
be present an intense blue ring will develop at 
the line of separation. 

This test is so delicate that it may detect blood 
even when the spectroscopic test is negative. Pus FlG - ^"^Jjj 11 crystals - 
will not lead to error if the solutions have been 

properly kept. A control should always be made with a fluid known to 
contain blood. While certain other bodies sometimes present may give a 
positive test yet a negative result always means that no blood is present. 

The most delicate of all tests for blood in the urine is the phenolphtha- 
lein test of Kastle. 141 This is said to show 8 parts of blood in 1,000,000 
parts of urine. 

Spectroscopic Tests. — The spectroscopic examination of a urine which 
contains blood will usually show a mixed spectrum. If the blood is fresh 
the lines of oxyhemoglobin will predominate, but in cases of hemoglobin- 
uria or of nephritis, those of methemoglobin mil be conspicuous. Bacteria 
will tend to oxidize this lack back to oxyhemoglobin. The urine used must 
be clear and should if necessary be diluted. 

Very small amounts of blood-pigment may be detected as follows 
(Hoppe-Seyler) : To 100 c.c. of the urine is added a solution of albumin or 
an albuminous urine. The urine is then coagulated by heat, filtered, the 
precipitate washed, pressed out and rubbed up with alcohol which contains 

141 Bull. 51, Hygienic Lab., Public Health and Marine Hospital Service. 
16 



242 CLINICAL DIAGNOSIS 

a little H2SO4. It is then warmed and filtered. The nitrate after the 
addition of NaOH and (NH 4 ) 2 S gives the spectrum bands of hematin. 

Methemoglobin. — Many previous reports of the presence of methemo- 
globin in the urine are not reliable since hematin was not excluded . Methe- 
moglobin is present in all fresh urines containing blood, although in time 
the bacteria may oxidize it to oxyhemoglobin. The spectrum of neutral 
methemoglobin resembles close] y that of hematin, but if ammonia be added 
that of alkaline methemoglobin will appear. This spectrum may be con- 
fused if other bodies which either have a spectrum of their own or which 
darken the field, as bile or urobilin, also are present. One must be careful 
not to dilute the urine too much for this test. 

Urobilin and bile-pigment may be removed from the urine with basic 
PbAc, in which case hemoglobin will remain in solution, but methemoglobin 
will be precipitated. In case the hemoglobin is still present in red 
blood-cells, water should be added in sufficient amount to lake them. If 
the spectrum of hemoglobin cannot be seen or is very faint the hemo- 
globin may be transformed to reduced hematin whose spectrum is much 
easier to study. This reduction is best accomplished with (NH 4 ) 2 S, or 
with NaHSO^ and zinc. If the action of these reagents is prolonged 
albumin will be precipitated. The spectrum of reduced hemoglobin is 
fainter than is that of oxyhemoglobin. 

Hematoporphyrin. — Hematoporphyrin, an iron-free derivative of he- 
moglobin, is present in normal urine in traces, but under certain conditions 
it may appear in very large amounts . The most important of these condi tions 
are : the protracted use of sulphonal especially, but also of trional and tet- 
ronal; the acute infectious diseases, including acute rheumatic fever and 
pericarditis; the various forms of tuberculosis especially; Addison's disease; 
paroxysmal hemoglobinuria ; pneumonia ; lead poisoning ; hematemesis ; while 
some claim that an increase of this pigment in the urine is important in 
the diagnosis of liver disease, especially of cirrhosis. Pal 142 reported a case 
of paroxysmal hematoporphyrinuria due, he thinks, to lues, with " black " 
urine and with symptoms similar to those of paroxysmal hemoglobinuria. 

Urines rich in hematoporphyrin usually have a color described as dark 
brownish-red, cherry-color, Bordeaux-red, or " Port-wine " color. This 
color is, however, sometimes deceptive, especially in the cases due to lead 
poisoning, since it is not the hematoporphyrin itself which explains the 
peculiar color. In fact, considerable of this pigment may be present in a 
normally colored urine, while all of the hematoporphyrin may be removed 
from a highly colored urine without changing its color and, finally, some 
urines whose color strongly suggests the presence of this pigment contain 
none at all. What the pigments are which are so often excreted with hema- 
toporphyrin is not known. 143 

142 Centralbl. f. inn. Med., 1903, vol. xxiv, p. 601. 

543 Monro, Quart. Jour. Med., 1907, vol. i, p. 49; 1910, vol. iv. 



THE URINE: SEDIMENTS 243 

About forty fatal cases of hematoporphyrinviria following the use of 
sulphonal have been reported. The most of these were women. Garrod 144 
collected 12 cases not due to sulphonal, nearly all in men whose condition 
lasted for years without any bad symptoms. 

The cause of hematoporphyrinuria is not known. Some claim that lead, 
sulphonal, etc., have a direct action on the red blood-cells; others, that they 
cause hemorrhages into the gastric mucosa and that the blood-pigment 
there liberated is changed by the gastric juice to hematoporphyrin which is 
absorbed and excreted ; while in other cases the trouble is said to lie in the 
renal epithelium. An hematoporphyrinemia has not yet been proved. 145 
Some ascribe the condition to perverted catabolism of hemoglobin rather 
than to increased destruction of red cells. 

Salkowski s Method of Detecting Hematoporphyrin in the Urine. — From 
30 to 50 c.c. of urine is completely precipitated by an alkaline solution of 
barium chloride (equal parts of a cold saturated solution of barium hydrox- 
ide and of a 10% solution of barium chloride) and filtered. The precipitate 
is washed, first with water and next with absolute alcohol. It is then 
extracted by pouring repeatedly warm acidified alcohol (10 c.c. of alcohol 
containing from 6 to 8 drops of HC1) over the precipitate on the filter paper. 
If hematoporphyrin is present the alcoholic filtrate will become reddish- 
violet in color, due to acid hematoporphyrin, and its spectrum will show 
two absorption bands. If now ammonia be added the color of the solution 
will change to yellow and the spectrum will show the 4 bands of alkaline 
hematoporphyrin (Sahli) . 

SEDIMENTS 

Preservation of the Urine. — To study organized sediments the urine 
should be examined while perfectly fresh, for casts often disintegrate 
rapidly. The best way to get the sediment is to allow the urine to stand 
in a conical glass in a room at low temperature. The most convenient 
way, however, is to centrifugalize the fresh urine in a centrifuge. One 
great disadvantage of this method is that the high pressure at the point 
of the tube may compress casts, cells and mucus into an indistinguishable 
mass. A quick but not accurate way is to filter the urine through a filter 
paper and examine the last few drops in the funnel. A drop of the sediment 
is drawn up from the point of the conical glass, centrifuge tube, etc., into 
a clean pipet; the outside of the pipet is wiped off and the drop blown 
onto a large, clean glass slide. No cover-glass is necessary for the prelimi- 
nary survey of the field by low magnification, but a large cover-glass should 
later be dropped over interesting positions of the field and the higher dry 
powers used. If the urine contains very much sediment this will settle in 
layers, the composition of which will vary considerably. Such a sediment 
should first be well mixed or the several layers separately examined. 

144 Lancet, March 5, 1904. 

145 Ruedy, Am. Jour. Insanity, October, 1899. 



244 CLINICAL DIAGNOSIS 

To preserve the urine from bacterial action during long sedimentation 
a piece of camphor or of thymol or a few drops of formalin are best, but 
the latter may add to the sediment a component of its own. Chloroform, 
so valuable in the preservation of crystals and for chemical work, cannot 
be used to preserve casts and cells. 

To preserve sediments for long periods of time the urine is centrifugal- 
ized and the supernatant fluid poured off. Chloroform " is then added 
to preserve crystals, or formalin to i to 2% to preserve casts and 
formed elements. 

Since there are very few inorganic sediments which can be recognized 
beyond doubt from their appearance alone the student should practise the 
methods of microcherristry. The reagents may be drawn under the cover 
by applying the edge of a piece of filter paper to the edge of the cover- 
glass opposite to a drop Of reagent used which just wets the edge of the 
cover-glass. 

There is one peculiarity of crystalline sediments worthy of mention, 
that the crystals of any one substance in any one urine all usually belong 
to the same system. Another peculiarity is the relative infrequency of 
crystalline sediments in women's urine. 

The sediments have been divided into those which are organized and 
unorganized. These terms are relics of an antiquated pathology. By the 
former are meant tissue elements, casts, bacteria, and formed elements 
from the urinary or communicating organs; by the latter, any precipitate. 
They may also be divided into gross and microscopic sediments. The 
normal urine when passed never has a gross sediment but always a micro- 
scopic organized sediment consisting of a trace of mucus and a few mono- 
nuclear cells. The uncatheterized specimen of a woman's urine will prac- 
tically always contain also a few cells from the genital tract which are 
washed by the stream of urine from the mouth of the urethra and the labia. 
After the urine has stood for even a few minutes crystals or a gross amor- 
phous sediment may appear, the amount, composition and character of 
which will depend on the temperature, the concentration and reaction of 
the urine and on the rapidity of ammonia production. It will depend 
much less on the diet. 

A gross sediment of phosphates and carbonates of the alkaline earths 
may cloud the urine when passed in cases of phosphaturia, persons whom 
we must consider normal. After a normal urine, perfectly clear when 
voided, has stood for a shorter or longer time, an inorganic precipitate, often 
abundant, may appear, of uric acid or urates in an acid urine and of phos- 
phates and carbonates in an alkaline urine. In pathological conditions the 
organized sediments often are gross. These may consist of blood, pus, 
cystin, even of casts (see page 272) while an inorganic alkaline phosphate 
sediment may be voided with the urine in various diseases of the kidney 
and urinary tract. 



THE URINE: SEDIMENTS 245 

The reaction of the urine may often be indicated by the appearance of 
a gross sediment; when acid, this is granular; when alkaline, it is mucoid. 
Preparations for microscopic study should be examined as soon as made, 
since if allowed to dry even a little the crystals or urea, NaCl, etc., which 
separate, may confuse one. 

Unorganized Sediments — (i) Urates and Uric Acid. — The urates 
may separate from any normal concentrated acid urine, especially on a 
cold day (much to the distress of mind of some persons) first as a remarkably 
muddy or milky cloud, then as a heavy voluminous sediment, the color of 
which will vary from a yellow to a bright rose-red, which settles on the 
bottom and sides of the container. It disappears at once on warming. 
This is the most characteristic reaction. It is soluble in acids, with the 
subsequent precipitation of uric acid, and in alkalies. This sediment is 
especially common in the urine of patients with fevers, e.g., pneumonia, 
acute rheumatic fever and in chronic passive congestion. It rarely forms 
in albuminous urines. 

This precipitate is said to be composed, of the quadriurates (Roberts), 
MHUU, which are said to be formed by the action of MH 2 P0 4 on the 
biurates, MHU. If present in sufficient concentration the quadriurates 
will precipitate as such, but if present in less-concentrated solution they 
will be decomposed, forming biurates and uric acid, the latter of which 
will at once precipitate as little, bright red, so-called " red pepper granules " 
on the sides of the glass. In the urate sediment are found also calcium 
oxalate crystals and, as ammonia soon forms, ammonium urate globules. 
Hence in the same sediment one may find ammonium urate, the so-called 
quadriurates, uric acid, calcium oxalate and even a few triple phosphate 
crystals. This transformation of the sediment progresses from above 
downward. 

This explanation of Roberts would be very satisfactory were it not that 
it has little evidence behind it. One thing is quite certain, that the pre- 
cipitation of the urates is the result of a chemical and not merely a physical 
change in the urine. The precipitate forms too slowly in the cooling urine 
to be due to temperature changes alone and warming the urine to its original 
temperature does not redissolve this sediment. Also, during its formation, 
the acidity of the urine is said to increase. 

Microscopically, the acid urate sediment consists of clusters of very 
fine granules, the color of which varies from yellow to a reddish -brown, 
which disappear on warming. The addition of a little acetic acid will be 
followed by the appearance of uric acid crystals. 

Ammonium Biurate. — Arnmonium biurate (see Fig. 37) is the only 
urate sediment which forms in an alkaline urine. It may indeed begin to 
appear while the urine is still very faintly acid as soon as enough ammonia 
is available, it will increase in direct proportion as ammonia is formed and 
later will be mixed with a sediment of amorphous phosphates and triple 



246 CLINICAL DIAGNOSIS 

phosphates. Microscopically it may consist of spheres which often present 
the so-called " morning star " or " thorn apple " shape, which have a dark 
color, are often concentric or radially striated and are covered with crystals 
projecting like thorns; but more often it assumes very irregular, bizarre 
shapes. This precipitate is soluble in acetic acid with the subsequent 
precipitation of uric acid and the liberation of ammonia. 

Uric acid and the urates when pure are colorless but their common 
yellow color is due to urochrome especially, but also to urobilin, and the 
red to uroerythrin. The precipitating urates have a peculiar affinity for 
bile pigments and so this sediment may carry down and contain all the 
bile that there is in a specimen of urine. The same may be true of the black 
pigment of the urine in cases of carbolic acid poisoning. 

The needle crystals of so- 
dium biurate are rarely found 
in the urine and then only in 
urines undergoing ammoniacal 
decomposition but which are 
still amphoteric. These crys- 
tals resemble calcium phos- 
phate, but when brought into 
contact with acetic acid they 
at once dissolve and a cloud of 
uric crystals appears. 

Uric Acid. — Uric acid (see 
Fig. 38) when pure crystallizes 
usually in rhombs, but in the 

F T G. 37. — Various forms of ammonium biurate crystals. urine rhombs with definite 

angles are rarely if ever seen 
for the angles are rounded and the crystals have the so-called "whet- 
stone " shape. When seen on the edge these crystals are very narrow 
rectangles. They may be single, in rosettes, or clustered in the shape of a 
barrel (see Fig. 38, a, b, 1). 

Sometimes the uric acid crystallizes in needles arranged in sheaves (see 
Fig. 38, 4). These crystals may occur as masses as large as the head of a 
pin, which cling to the glass (see Fig. 38, 2). Their color varies from yellow 
to brown, or they may be colorless. The colorless crystals are sometimes 
perfect hexagons in which case their recognition is difficult, since they may 
in appearance perfectly resemble cystin crystals. Dr. T. B. Futcher once 
provided our class in clinical diagnosis a remarkable specimen of urine 
which illustrates this point. The patient, a girl six years of age with dia- 
betes mellitus, had been voiding from 1000 to 2000 c.c. of urine per day, 
with a specific gravity about 1.035; sugar, from 5.1 to 5.5% and nothing 
of interest microscopically. One day after 24 hours on a carbohydrate-free, 
proteid-rich diet the urine (its sp. gr. was 1.026 and the sugar, 0.6%) was 




THE URINE: SEDIMENTS 



247 



turbid because of a suspension of glistening, colorless, hexagonal crystals 
which in appearance exactly resembled cystin. Many were single, but 
most were in clumps of even macroscopic size. It was only on chemical 
examination that they could be recognized as uric acid crystals. On this 
day no typical uric acid crystals were seen. The following day the sediment 
consisted of a mixture of hexagonal and whetstone crystals and after that 
not a single hexagonal form was found. 

The yellow color of uric acid crystals is due to urochrome, not to uro- 
bilin, and the red to uroerythrin plus urochrome. Hematoporphyrin, 
bilirubin, or biliverdin may if present give their color to these crystals. 
In cases of carbolic acid poisoning they may have a dark brown, almost 
black, color. 

Crystals which are precipitated artificially by the addition of an acid 
to a urine have a reddish-brown 
color due to black decomposition 
products of urochrome ; their 
color may also be due to indigo- 
blue or indigo-red, if these are 
present in the urine. 

Calcium urate crystals are 
said to appear sometimes in the 
same sediments with calcium 
oxalate crystals. They are color- 
less prisms, insoluble in hot water, 
give the murexid test and are 
soluble in acid with the subse- 
quent appearance of uric acid crystals. They may be produced by 
treating an acid urate sediment with lime water. 

Detection. — The urate sediments which are deposited in acid urine 
may usually be recognized from their gross appearance alone, but the char- 
acteristic tests are that they disappear on warming and that they all are 
dissolved by acid with the subsequent precipitation of uric acid. Uric acid 
crystals are not dissolved by heat or by acid. Ammonium biurate spheres 
are characteristic in form and are soluble in acid followed by the appear- 
ance of the uric acid crystals. 

Murexid Test. — This test is characteristic of uric acid and its 
salts. The crystal or sediment to be tested is evaporated in a porce- 
lain dish with dilute HNO3, and to the residue is added some weak 
NH4OH. If uric acid (or urates) is present a beautiful purple-red color 
will appear. 

A urate sediment has little significance except that it indicates a con- 
centrated acid urine. A uric acid sediment, however, may have great 
importance since it sometimes forms large concretions in the pelvis of the 
kidney or bladder. 




Fig. 38. — Uric acid crystals. (The lettered forms are 
drawn from nature, those numbered are copied from 
Rieder's Atlas.) 



248 CLINICAL DIAGNOSIS 

Phosphates and Carbonates. — (i) Amorphous earthy phosphates 
and carbonates may be precipitated in any urine by the addition of a 
little fixed alkali. A somewhat similar precipitate forms when a weakly 
acid or alkaline urine is heated, since the acid salts of phosphoric and car- 
bonic acid are then changed to insoluble basic salts. Both are soluble in 
acetic acid, the carbonates with gas evolution. They are the chief constit- 
uent of the sediment of an alkaline urine and may cloud even the fresh 
urine of certain nervous cases and of cases of gastric hypersecretion who 
lose much acid from the stomach because of vomiting, lavage, or diarrhea. 
The total amount of phosphoric acid in the urine of these cases of so-called 
phosphaturia is not increased. Microscopically this precipitate is seen to 
consist of very coarse colorless granules varying considerably in size, which 
disappear on the addition of a little acetic acid. By the gas bubble formed 
one can tell which granules were the carbonates. 

(2) Triple Phosphates, MgNH 4 P0 4 6H 2 0. — The beautiful triple phos- 




Fig. 39. — Various forms of triple phosphate crystals. Fig. 40. — Atypical forms of triple 

X 400. To the left are coffin-lid shapes; in the lower phosphate crystals. X 400. 

center a perfect pyramid; that in the upper left corner 
resembles neutral magnesium phosphate; that in the 
upper right is a partially dissolved crystal. 

phate crystals (see Fig. 39) appear in urine even while still acid as soon as 
sufficient ammonia is present. They appear usually in connection with the 
amorphous carbonates and phosphates, often with ammonium urate, while 
in some urines they are the chief constituent of the sediment. These 
crystals belong to the rhombic system. In size they vary from those very 
small to some even 9 mm. in length. The so-called coffin-lid crystals 
are characteristic (see Fig. 39), but many modifications of this shape may 
be found. Other strange X-forms are due to the partial solution of a 
crystal. Some are said to resemble calcium oxalate crystals but this 
resemblance is only superficial, for even when triple phosphate appears 
as a perfect pyramid with a square base there is no trace of a double 
envelope appearance (see Fig. 39 and page 251). 

Fern-shaped crystals occur especially in sediments artificially precipitated . 

In some urines nearly all the triple phosphate crystals have unusual 
shapes; some are very thin plates (see Fig. 39); some have bevelled edges, 
some apparently not ; some have square, others rounded or bevelled corners, 



THE URINE: SEDIMENTS 249 

some are wedges (see Fig. 40), some triangular prisms; yet all have a 
greenish hue which is not seen in the calcium oxalate crystals. 

Neutral Magnesium Phosphate, Mg 3 (P0 4 )222H 2 0. — The very rare 
crystals of neutral magnesium phosphate (see Fig. 47, b) have been found 
in alkaline urines in which the amount of ammonia was not sufficient to 



Fig. 41.— Wedges of dicalcium phosphate. Fig. 42.— Sheaves of calcium phosphate needles. 

X 50. 

form triple phosphates. Such happens in certain cases of dilated stomach 
with considerable vomiting and also after the ingestion of magnesium 
carbonate, etc. These crystals are exceedingly refractile, long rhombic 
tablets with bevelled edges. Some resemble the very thin coffin-lid triple 
phosphate crystals (see Fig. 39). This is a beautiful sediment to study. 

Dicalcium Phosphate. — Di- 
calcium phosphate crystals are 
sometimes but rarely found in 
amphoteric or weakly acid urines. 
They are small prisms or wedges 
arranged in irregular clumps (see 
Fig. 41), or massed together in ro- 
settes (see Fig. 47, d) or fan-shaped 
clusters . In these masses or rosettes 
the individual small crystal can 
hardly be made out. A rather un- 
usual form of calcium phosphate 
crystal is shown in Fig. 43 and a 
still more unusual form in Fig. 42. Dicalcium phosphate may crystallize out 
when the urine is rich in calcium and only weakly acid, which would seem 
to occur especially in chronic arthritis. They are soluble in acetic 
acid. They may be distinguished from triple phosphates crystals since 
20% ammonium carbonate will dissolve the dicalcium crystals and not 
the latter. 

Calcium Carbonate. — These crystals (see Fig. 44) may be mingled 
with the amorphous carbonates in an alkaline urine. They occur as 
amorphous masses or as dumb-bells resembling somewhat the CaOx 




Fig. 43. — Calcium phosphate (?). X 400. 



250 CLINICAL DIAGNOSIS 

crystals, or as large concentric radiating spheres. They are soluble in 
acetic acid with gas liberation. 

Neutral calcium phosphate may appear as a scum on the surface of 
even quite fresh urine, resembling a film of oil which may easily be skimmed 
off. This scum under the microscope is seen to consist of amorphous 
matter in sheets. This precipitate often is a nuisance since it clings to the 
outside of a pipet used for obtaining samples of the sediment. 

Oxaluria. — The symptom complex, oxaluria, formerly so respected, is 
seldom mentioned now. This term was used of any nervous condition if 
the urine of that patient contained a large sediment of CaOx crystals. 
The precipitation of these crystals, however, does not depend as much on 
the total amount of oxalic acid present as it does on its solubility. The 
precipitation of CaOx in the urinary tract, on the other hand, is of very 
great importance since from 30 to 50% of urinary calculi consist of CaOx 
and these are the worst of stones.. 

While a certain amount of the CaOx of the urine is a product of tissue 
combustion, since some is present in the urine even of a starving person, 

, yet the chief source is the food, especially certain 

\ vegetables as beans, artichokes, beets, potatoes and 

^jfy . especially tomatoes, spinach, rhubarb, certain fruits 

and grains, cocoa, tea and coffee. Only about 15% 

of the oxalic acid ingested is absorbed into the blood- 

Fig. 44.— Calcium carbon- stream and this is excreted quantitatively in the 

ate. X 400. 

urine as CaOx; about 10% of that ingested appears 
in the stools ; the rest is destroyed by the intestinal bacteria and ferments. 

In health the output in the urine averages about 20 mgms. per day; 
the upper limit, 35 mgms. Bakhoven thinks that the carbohydrates of 
the foods are the chief oxalate builders. In its excretion oxalic acid 
bears no relation to the uric acid output; the latter, for instance, can be 
increased by a diet rich in the nucleins and there be no change in the 
oxalic acid output. 

Calcium oxalate crystals, present in very fresh urines, attracted con- 
siderable attention of the older pathologists, as they were supposed to 
cause an irritation in the urethra and so explained many of the symptoms 
and vicious habits of neurotic individuals, e.g., masturbation. 

Among the diseases claimed to be accompanied by oxaluria are pulmon- 
ary tuberculosis, peritoneal tuberculosis, pernicious anemia, leukemia (in 
which condition, it is claimed, the output varies from 33.2 to 53 mgms. per 
day), jaundice, diabetes mellitus, gout, diseases of the digestive and respira- 
tory organs, cirrhosis of the liver and, especially, neurasthenia. The 
amount of oxalic acid in the urine bears some relation to the absence of 
HC1 in the gastric juice and to fermentation processes in the intestine. In 
diabetes mellitus a large output is usually present, which increases as the 
sugar diminishes (vicarious oxaluria). Naunyn mentions three cases in 



THE URINE: SEDIMENTS 



251 




Pig. 45. — Various forms of calcium 
oxalate crystals and spheres. X 400. 



which quantitative estimates of CaOx were made. One eliminated 0.8 gm., 
the second 1.2 gms. in 24 hours and the third 0.5 gm. per liter. 

It is of interest that some insurance companies regard oxaluria as an 
early sign of nephritis. The best recent work on this subject is that of 
Serkowski and Mozdzenski 146 who showed by accurate methods that there 
is no apparent relationship between the amount of oxalic acid in solution 
and that in the sediment, or between the 
amount of oxalic acid and of uric acid in 
the urine. Its output does, however, run 
roughly parallel to that of the acid phos- 
phates. 

Calcium oxalate crystals may precipi- 
tate in any urine but the real question 
chemically is, Why does any remain in 
solution? Klemperer and Tritschler 147 con- 
sider that the acid phosphates aid in 
holding it in solution, those of sodium 
least, those of calcium more and those of magnesium most; also that the 
absolute amount of CaOx in the urine is of importance. It is almost 
impossible to associate this sediment with any pathological condition. 
CaOx crystals occur in 2 forms, the first of which, the double envelope 
octahedral form, is quite characteristic. On a hasty glance it may 
■ resemble some square triple phosphate crystals, 
but they are insoluble in acetic acid. The second 
form, the spherical, could be mistaken on hasty 
inspection for CaC0 3 crystals, although they show 
a different structure and are insoluble in acid. 

(1) The octahedral (CaC 2 043H 2 0) which be- 
long to the tetragonal system (see Fig. 45) resemble 
double envelopes or prisms. 

(2) The spheroidal forms (CaC 2 044H 2 0) (see 
Fig. 45) are flat, oval, or nearly semicircular with a central groove and 
hence resemble an hour-glass. They often show a radial striation. 

A rate form of oxalate crystal is represented in Fig. 46. These crystals 
were flat plates with parallel sides and rounded ends and resembled super- 
imposed sheets of mica. 

Calcium oxalate crystals are transparent, very refractive and usually 
colorless, but may be bile-stained if the urine contains bile pigment. They 
are insoluble in water, are very little soluble, if at all, in acetic acid, and are 
easily in any mineral acid. As this precipitate forms very slowly the crystals 
forms are perfect. They may be found in acid, amphoteric, or weakly 
alkaline urine, and are sometimes present in the specimen when voidedo 



V- : 



)J 



Fig. 46. — -A rare form- of 
calcium oxalate crystals. 



146 Zeitschr. f. phys. Chem., Jan., 191 1, vol. lxx, p. 264. 

147 Zeitschr. f. klin. Med., 1902, vol. xliv, p. 337. 



252 CLINICAL DIAGNOSIS 

Quantitative Determination of Oxalic Acid. — The Neumann 
method is as follows: The 24 hours' collection of urine is precipitated with 
calcium chloride and ammonia and then made weakly acid with acetic acid. 
A small amount of alcoholic thymol solution is added to inhibit bacterial 
growth. The urine is now allowed to stand for over 24 hours in a warm 
place. The precipitate is then washed several times by decanting the super- 
natant fluid through a filter paper and finally bringing the entire precipitate 
onto the paper. As much of the washing as possible is done by decanting., 
since the fine precipitate passes easily through the paper. The precipitate 
is then dissolved in warm dilute HC1, and the paper washed with water 
until the water is no longer acid. This filtrate is evaporated to a small 
volume in a porcelain dish on the water -bath, then poured into a small 
stout cylinder, washing the dish with water and dilute HC1 and adding 







■ >; 


■.fp : 


p- -..._ 


" V . 






^-, 


' C^'~ :jJ 


1 .;-.■ 


a 


"V; 




hi c 


, d 






• v ) 

e - 


N c .-.■; 



Fig. 47. — Various crystals: a, calcium sulphate; b, neutral magnesium 
phosphate; c, hippuric acid; d, acid calcium phosphate; e, colorless 
uric acid. (Copied from Rieder's Atlas.) 

the washwater to the fluid. Ammonia is then added in excess, a few drops 
of litmus being added to assure one of the reaction. After at least 24 hours 
standing, the precipitate is brought onto an ashless filter paper, loosening 
those crystals which cling to the walls of the cylinder with a glass rod the 
end of which is covered with a small piece of rubber tubing. The precipi- 
tate is then washed with water, until it is chlorine-free, and then with acetic 
acid. The filter is now dried, burned in a platinum crucible first at a dull 
red then with a blast flame until at constant weight. Calcium oxalate is 
thus transformed to calcium oxide, 50 parts of which correspond to 90 parts 
of oxalic acid. 

Calcium Sulphate (CaS0 4 2H 2 0). — Calcium sulphate forms a very rare 
sediment found only in very acid urines. The crystals (see Fig. 47, a) are 
long, thin tablets or needles, some single but the most in clusters, which 
are insoluble in NH4OH, alcohol, and acetic acid and soluble with difficulty 
in HC1, HNO 3 and in hot water (little in cold) . The nature of these crystals 
should be confirmed chemically by dissolving them and proving the presence 
of sulphuric acid by the addition of a solution of BaCls. 



THE URINE: SEDIMENTS 



253 



Hippuric Acid. — Hippuric acid occurs rarely as a sediment of milk-white, semi- 
transparent, 4-sided prisms and rods with ends of 2 to 4 planes (see Fig. 47, c). These 
are distinguished from uric acid, which they may resemble in form, by their greater 
solubility in water, especially in warm water, their solubility in alcohol and ether and 
in that they do not give the murexid test. The amount of hippuric acid in the normal 
urine varies from o. 1 to 1 gm. per day, depending on the diet. 

Hetero-albumose, Bence- Jones Body. — In 2 cases Bence-Jones body, once crystal- 
line and once in amorphous condition, has formed a urine sediment. 

Xanthin. — Two or three instances have been reported with xanthin 
crystals in the urine sediment. These crystals resembled uric acid some- 
what (see Fig. 48, d) but are soluble on heating and in ammonia. If evapor- 
ated on a bath in quite concentrated NHO3 they give a yellow residue, 
which color on further careful heating over a small flame becomes more 
intense. If now KOH is added the color becomes first yellowish-red, then 
a deeper red or even a violet-red. 
This test should not be confused 
with the murexid test. 

Hematoidin (Bilirubin). — 
Hematoidin may appear as 
needles (Fig. 48, a) or rhombs 
(b) or in amorphous form in 
the urine sediment in hemor- 
rhagic nephritis, in very jaun- 
diced urine especially if acid has 
been added, in acute yellow at- 
rophy, in pyonephrosis, with 
cancer fragments and after trans- 
fusion. In cases of jaundice of the new-born they may be found in the 
epithelial cells in the urine. They have also been found in connection 
with waxy kidney, scarlet fever, typhoid fever and carcinoma of the liver 
with jaundice. 

Indigo. — Crystals of indigo, as a scum of blue needles arranged in stars 
or of blue rhombic plates which are soluble in chloroform giving a blue 
solution, may be found in normal urine undergoing decomposition, in the 
urine in peritonitis, pyelo-nephritis, etc. One also sees bundles of violet-red 
crystals or plates of indigo-red. 

Melanin as amorphous scales has been found rarely in the sediment. 

Hemoglobin in amorphous scales, plates, or casts occurs in the urine 
sediment in cases of hemoglobinuria. 

Cholesterol. — Cholesterol, as flat superimposed plates often with re-en- 
trant angles (see Fig. 22), is sometimes present in the urine in such amounts 
as to justify the term " cholesterinuria." It is found, always with other 
fats also, where large numbers of pus cells are undergoing fatty degeneration 
and when tissue is breaking down, therefore in some cases of vesical catarrh, 
in pyelitis especially, in pyonephrosis, echinococcus cysts of the kidney 




Pig. 48. — Various crystals of the urine: a, hematoidin 

needles; b, hematoidin crystals; c, leucin; d, xanthin; 

e, tyrosin. (Copied from various authors.) 



254 CLINICAL DIAGNOSIS 

and sometimes in nephritis. It is rarely found in the tissue of kidneys 
which have undergone fatty degeneration and never in chyluria, in which 
condition especially one would expect to find it. Hirschlaff 148 determined 
even 5.8 gms. of cholesterol per 100 c.c. of urine in the urine of a case of 
hydronephrosis while the sac was emptying. We followed for some time 
a marked case of cholesterinuria of long standing in a case of renal cyst 
of doubtful nature. 

Cholesterol is insoluble in cold, but easily in hot, alcohol, the precipitate 
reappearing as the alcohol cools. It is soluble in chloroform. If a solution 
of cholesterol be superimposed on concentrated H2SO4 its color becomes 
first blood-red and then more violet -red, while that of the sulphuric acid 
becomes dark red with a green fluorescence (Salkowski). This play of 
colors can be watched under the microscope if cholesterol crystals be 
brought into contact with H 2 S0 4 4 parts and H 2 1 part. 

Leucin and Tyrosin. — Leucin and tyrosin appear in the urine in certain 
pathological conditions. Leucin is never a spontaneous sediment while 
tyrosin needles have been reported as a sediment in 3 cases, 1 of which was 
of acute yellow atrophy of the liver, and 1 of phosphorus poisoning. These 
substances in solution are often present in the urine. of cases with acute 
yellow atrophy, phosphorus poisoning, smallpox (rarely), severe typhoid 
fever, pernicious anemia, leukemia and sometimes in cases with simple 
cardiac liver. 149 To demonstrate them it is as a rule necessary to evaporate 
the urine to about Xo its volume and then add alcohol which will precipitate 
the needles of tyrosin and the spheres of leucin (as well as peptone and 
lactic acid). Tyrosin crystallized out in black needles grouped together 
like sheaves of wheat (see Fig. 48, e). Since patients with tyrosin in the 
urine are usually jaundiced one may confuse tyrosin with the intense brown 
needles of bilirubin which often have a rather similar shape. In an alkaline 
urine the calcium phosphate needles must be excluded. Leucin should be 
searched for in fresh urine since it may form rapidly in any decomposing 
albuminous urine. If pure it would appear in groups of spherules (see 
Fig. 48, c) which have little refractility, a much clearer contour than the 
spherules of the urates, no spicules, and a hyaline or a radiating structure. 
But as a sediment it is practically always impure and in dark spheres or 
masses or as spherules which may have a dark center and a clear periphery, 
or vice versa. 

The presence of leucin in a sediment should always be confirmed by 
chemical tests. 

To isolate tyrosin and leucin from urine it is necessary, first, to remove 
all albumin by heat and acid. The filtrate is first precipitated with neutral, 
then with basic, lead acetate until all precipitation ceases. The urine is 
then filtered, the lead in the filtrate removed with H 2 S and the filtrate 

148 Deutsches Arch. f. klin. Med., 1899, vol. lxii, p. 531. 

149 Mann, Quart. Jour. Med., October, 1907. 



THE URINE: SEDIMENTS 255 

concentrated by evaporation. The tyrosin may begin even now to separate 
out slowly if present in considerable amount. The fluid should be con- 
centrated to very small volume and the urea extracted by absolute alcohol. 
The residue is then boiled with weak ammoniacal alcohol and the filtrate 
is again evaporated to small volume and then allowed to crystallize. The 
leucin and tyrosin will separate out, that one first the solution of which 
first reaches saturation. A partial separation may now be accomplished 
by adding a small volume of alcohol which will dissolve the leucin more 
easily than the tyrosin. If after the above procedures no precipitate has 
appeared the residue and fluid are again diluted, precipitated with basic 
lead acetate and the process repeated. 

The leucin and tyrosin could be more completely separated by dissolving 
the residue after evaporation in boiling water plus a little ammonia and 
adding to the hot solution basic lead acetate, stirring all the while, until 
the precipitate is no longer brown but white. This is then filtered, heated 
nearly to boiling, made slightly acid with dilute H 2 S0 4 , and then boiled 
to drive off the ammonia and to precipitate the lead. It is then rapidly 
filtered and cooled. The tyrosin will now precipitate out almost quanti- 
tatively. The solution remaining is now heated with H 2 S to precipitate 
the lead and then evaporated to smaller volume. While it is boiling freshly 
precipitated Cu(OH) 2 is added in excess and the boiling continued for a 
few minutes. The precipitate, which will contain part of the leucin, is 
suspended in boiling water, decomposed with H 2 S, a little acetic acid added 
and then filtered. The filtrate is decolorized with animal charcoal and 
evaporated to small volume. On cooling, some of the leucin will now 
separate out and the rest will remain a blue copper compound. It is 
very difficult to obtain leucin pure although this can be done by forming 
its ethyl ester. 

Tyrosin (C 6 H 4 CH 2 CHNH 2 COOH).— Tyrosin (see Fig. 48, e) will 
crystallize out from aqueous solutions in bundles of needles arranged like 
sheaves of wheat and from ammoniacal alcohol solution in bunches of 
prisms. These crystals are soluble in water, slightly so in alcohol, not at 
all in ether and are easily in acids and alkalies. Tyrosin cannot be positively 
identified from the appearance of its crystals. Chemical identification is 
necessary, yet these tests cannot be applied directly to a urine or to a mixed 
sediment. The sediment should be filtered out, washed with water, dis- 
solved in ammonia plus a little ammonium carbonate in warm solution 
and then evaporated until it recrystallizes. 

Piria's Test. — To some dry tyrosin in a test-tube are added a few drops 
of concentrated H 2 S0 4 , this at first warmed gently and then boiled in a 
water-bath for % hour. A red solution of tyrosin sulphate is formed. This 
is now cooled, several volumes of water added, is then neutralized with 
BaC0 3 and filtered. The filtrate is evaporated to a few cubic centimeters 
and weak, acid-free Fe 2 Cl6 added to the cooled solution. A fine violet 



256 CLINICAL DIAGNOSIS 

color results. This test will be interf erred with by any free mineral acids 
or by an excess of Fe 2 Cl6. 

The hot aqueous solution of tyrosin gives with Millon's reagent, 
(Hg(N0 3 ) 2 +KN0 2 ), a fine red color and an abundant red precipitate. 

Leucin [(CH 3 )2CHCH 2 CHNH 2 COOH].— Leucin will separate out from 
a solution as spherules whose color and regularity of outline will depend 
on the purity of the specimen. The spherules often separate out in groups 
and frequently show a striation. Leucin (see Fig. 48, c) is soluble in water, 
less so in alcohol, but very easily in acids and alkalies. All of these com- 
pounds are more soluble in an impure than in a pure condition. No leucin 
is ever found in a urine sediment before the urine has been much concen- 
trated. The methods for its isolation have been mentioned above. Before 
the chemical tests are applied leucin must first be purified by recrystallizing 
it from hot ammoniacal alcohol. Its characteristic tests are: Its crystal 
form when pure, the fact that it sublimes to a woolly mass at a gentle heat 
at 170 C. with fusion and with the odor of amylamine, Scherer's test and 
Salkowski's test. 

Scherer's Test. — Pure leucin together with a little HNO3 is evaporated 
on a platinum foil. A colored residue is obtained. This is warmed with 
NaOH and a water-clear if pure, or colored if impure, fluid results. If 
this is evaporated carefully an oily fluid is obtained which rolls around 
without wetting the foil. This test is characteristic for even impure leucin. 

Salkowski's Test. — To the specimen is added a little water plus 1 or 2 
drops of 10% CuS0 4 . A blue solution is obtained, (C*Hi 2 N0 2 )2Cu, which 
does not reduce on heating. 

Cystin. — Cystin, supposed to be a product of intermediate proteid 
metabolism which because of some perversion of metabolism jiot yet well 
understood escapes further cleavage, may appear for years, even for the 
individual's entire life, in the urine in large amounts. Cystinuria itself 
gives rise to no symptoms but the calculi it forms make the lives of their 
victims miserable and lead to repeated operations. Fortunately the forma- 
tion of calculi is intermittent, allowing the patients long periods of relief 
while in some cases the removal of the stones would seem to be followed by 
the cessation of the cystinuria. The output of cystin in some cases is said 
to be intermittent. 

Cystin is a normal intermediate product of proteid metabolism and 
contains the most, possibly all, of the sulphur of the proteid molecule. 
It is not a constituent of normal urine. If a normal person ingests cystin 
he will eliminate about 66% of its sulphur as inorganic sulphates and about 
/3 as neutral sulphur, but none as cystin. Simon and Campbell 15 ° think 
that some is eliminated in the bile as taurochloric acid. Why it should be 
excreted is not known. The theory that it is the product of an intestinal 
mycosis, which theory is borne out by the presence in both the intestinal 

150 Johns Hopkins Hosp. Bull., 1904. 



THE URINE: SEDIMENTS 257 

contents and urine of certain diamines, as cadaverin, putrescin and others, 
is no longer held. Most believe that it is an individual variation in metabol- 
ism, a congenital inability on the part of the organism to oxidize the cystin 
nucleus which may be present in several of the same family. 151 

In the fresh urine of a case of cystinuria may be seen the transparent 
hexagonal crystals of cystin. These crystals (see Fig. 49) are quite char- 
acteristic, yet not absolutely so since uric acid may assume this exact form. 
Sometimes these crystals are massed into large concretions from a pin-head 
to 1 cm. in diameter, of a whitish-yellow color, rather soft and waxy and 
crystalline on cross-section. These crystals are soluble in ammonia and 
reprecipitated by acetic acid, a test necessary to exclude uric acid. We 
have seen but 4 cases. One has had many of these concretions removed by 
crushing. Another case, a woman, was distressed for years by these con- 
cretions but refused operation. As she has since attained considerable 

success in public life we presume that the , 

concretions no longer bother her much. 

The urine in such cases often gives on 
standing the odor of H 2 S. It is in this condi- 
tion particularly that the neutral sulphur of the 
urine is greatly increased. The neutral sulphur 
is indeed the best index we have of the amount 
of cystin present. 

Diamines. — The presence of traces of diam- 
ines in the urine and in the feces has attracted l, 




some attention. 1 52 These found are putrescin Pig. 49— Cystin crystals from 

., , . . . urine. X 400. 

sometimes, cadaverin sometimes, sometimes 

both. Their presence is variable and intermittent. Lewis and Simon 

state in 1902 that they had been found in 7 cases. 

Baumann's method for their detection is as follows : The 2 4 hour amount 
of urine is shaken up with 10% NaOH and benzoylchloride (in the propor- 
tion of 1500 : 200 : 25) until the odor of benzoylchloride is gone. 

The precipitate (of phosphates, carbohydrates, and benzoylated diam- 
ines) is filtered with the aid of a suction-pump. The precipitate is digested 
with alcohol, filtered, the extract evaporated to small volume, 30 volumes 
of water added and then allowed to stand for from 12 to 48 hours. The 
benzoylated diamines will now separate out in the milky fluid as a volumin- 
ous sediment of white crystals. This is redissolved in alcohol, concentrated 
to small volume and diluted again with water. This process is repeated 
several times to separate the carbohydrates. 

More of the diamines may be recovered from the first nitrate by acidi- 
fying it with H2SO4 and extracting it 3 times with ether. To the ether 



151 Kretschmer, The Urologic and Cutaneous Review, 1916, xx, No. 1. 

152 Simon, Am. Jour. Med. Sci., 1900, vol. cxix, p. 39; 1902, vol. cxxiii, p. 838; Scholl- 
berg and Garrod, Lancet, August 24, 1901. 

17 



258 CLINICAL DIAGNOSIS 

residue 12% NaOH is added till the fluid is neutral, then 3 to 4 volumes 
of the alkali. If this is then kept in a cold place cystin and the diamines 
will separate out. They are filtered out and suspended in cold water; the 
benzoylchloride crystals remain. 

The crystals are dissolved in a little warm alcohol and 20 volumes of 
ether added. Benzoylputrescin will be precipitated. Its melting point is 
from 175 to 17 6° C. The ether residue contains benzoylcadaverin the 
melting point of which is 129 to 130 C. 

Unorganized Sediments. — The following outline given by Neubauer and Vogel for 
recognizing an unorganized sediment is so useful that we quote it in full. 

A. Acid urine. 

(a) Sediment amorphous. 

(1) Sediment consists of fine granules in clumps, mingled with which 

are crystals of uric acid and of calcium oxalate; urate sediment. 
This sediment is soluble on warming, and if a drop of strong 
acetic acid be added the granules gradually disappear with the 
subsequent separation in a few hours of uric acid crystals. 

(2) Dumb-bell shaped bodies. 

(a') Insoluble in strong acetic acid and soluble in concentrated 
hydrochloric acid without subsequent crystallization; cal- 
cium oxalate. 

(&') Insoluble in concentrate hydrochloric acid : probably calcium 
sulphate. This sediment should be filtered, washed, dis- 
solved in much hot water and tested for calcium and sul- 
phuric acid separately. 

(3) Very refractive globules, soluble in ether: fat. 

(4) Amorphous yellow granular masses: bilirubin or hematoidin. 

(b) Sediment crystalline. 

(1) Yellow or brown whetstone-shaped crystals, single or in rosettes; 

alone, or with amorphous urates and calcium oxalate: uric 
acid. These crystals are soluble in sodium hydroxide and repre- 
cipitated by an excess of concentrated hydrochloric acid. 

(2) Small yellow rhombic tablets alone or with amorphous granular 

tablets of the same color, often embedded in tissue detritus: 
bilirubin or hematoidin. 

(3) Colorless (or yellow in a decomposed urine), transparent, strongly 

refractive octahedrons, double envelope forms, or quadrangular 
short narrow prisms with octahedrons at the ends, insoluble in 
acetic acid and soluble in hydrochloric acid: calcium oxalate. 

(4) Crystals somewhat similar to the last mentioned, or large coffin-lid 

crystals, soluble in acetic acid: ammonium magnesium phos- 
phate (triple phosphates). 

(5) Symmetrical hexagonal tablets, sides and angle almost equal, insol- 

uble in acetic acid, soluble in ammonia: cystin. 

(6) Colorless whetstone-shaped tablets, insoluble in acetic acid; soluble 

in ammonia. On the addition of hydrochloric acid to this 
sediment in solution hexagonal tablets separate: xanthin. 

(7) Large, flat, strongly refractive elongated rhombic tablets, soluble 

in acetic acid, and partially so in ammonium carbonate: normal 
magnesium phosphate. 



THE URINE: SEDIMENTS 259 

(8) Prisms, single or in rosettes. 

(a') Soluble in ammonia: hippuric acid. 

(b') Insoluble in ammonia and in acids: calcium sulphate. 

(9) Wedge-shaped prisms, single, or in clusters or thick rosettes, decom- 

posed by ammonium carbonate and soluble in acetic acid: 
monacid calcium phosphate. 
(10) Bunches of very fine needles insoluble in acetic acid and soluble in 
ammonia and hydrochloric acid: tyrosin. 

B. The urine alkaline when the crystal precipitates (after the urine becomes alkaline 
many of the sediments previously mentioned which separated in the acid 
urine may still remain). 

(a) Amorphous. 

(1) Small granules together with triple phosphate crystals. 

(a') Soluble in acetic acid without gas formation: normal phos- 
phates of the alkaline earths. 

(£') Soluble, but with gas formation: carbonates of the alkaline 
earths. 

(2) Dumb-bell shaped masses or large spheres, soluble in acetic acid 

with gas formation: calcium carbonate. 

(3) Large dark spheres often covered by small projecting crystals: 

ammonium urate, soluble in hydrochloric acid or acetic acid 
with the subsequent separation of the rhombic crystals of 
uric acid. 

(b) Crystalline. 

(1) Large colorless prisms, many coffin-lid in shape: triple phosphates, 

easily soluble in acetic acid. 

(2) Rosettes of very fine blue needles or blue tablets : indigo. 

(3) Rosettes of violet-red needles or rhombic platelets: indigo-red. 

Chyluria. — Chyhiria is a condition characterized by the presence in 
the urine of enough fat in emulsion to give the urine a milky appearance. 
When the amount of fat is sufficient to give only an opalescent appearance 
the term lipuria is used, although the latter term should include both. 

There are 2 forms of chyluria, the 1 due to filaria infection, and the 
non-parasitic form, the etiology of which is not understood. Some claim 
that in the latter cases the source of the fat cannot be lymph as it is in the 
parasitic form since the urine contains no sugar as does lymph and also 
since there is often a higher percentage of fat in the urine than in lymph. 
Again, there is no decrease in the percentage of the normal urine constitu- 
ents as would be the case were the urine diluted by another fluid. 

In chyluria the fat may form gross tallow-like masses, but as a rule the 
droplets are even much finer than those of milk. The urine may resemble 
milk, in other cases, whey, but often it has a reddish tinge due to blood. 
The fresh urine is weakly acid or neutral in reaction and does not have the 
usual urinary odor. On standing a cream sometimes rises and a fibrin 
coagulum often forms. In addition to fat such urine always contains serum 
globulin and serum albumin and sometimes cholestrol and lecithin. Fibrin- 
ogen has been found, also hemialbumose and peptone. The amount of 



260 CLINICAL DIAGNOSIS 

proteid present varies from 0.2 to 2% or more and the fat from a trace to 
3%. A few leucocytes and a few red blood-cells may be found. In both 
forms urinary casts, etc., are absent unless Bright 's disease also is present. 
The urine may be chylous during the night and clear during the day, or 
vice versa. In other cases the excretion of the fat is intermittent, appearing 
only when the patient is in a vertical position, after digestion, after bodily 
exercise, or after excitement, etc. In some cases coagula form: in the bladder 
and cause considerable trouble. In parasitic chyluria one finds also in the 
urine, usually in the coagula, the eggs and embryos of filaria. Parasitic 
chyluria is a disease which lasts, often with intermissions, from months 
to years. It may cease spontaneously. This disease is endemic in certain 
tropical and subtropical regions, while a few cases are met with in the 
temperate zone. In these cases all the fat would seem to come directly 
from the pelvic lymphatics. 

In some cases a fat diet increases a chyluria and a foreign fat may even 
be recognized in the urine. Chyluria would seem not to be due to a renal 
lesion. Claude Bernhard held that it was due to hyperlipemia and this to 
the inability of the body to burn fat ; but an hyperlipemia is rare and this 
alone would not explain the albuminuria also present. Others ascribe 
chyluria to liver disease. Franz and Styskal believe that the fat comes 
directly from the lymphatic vessels since the chyluria will diminish or 
disappear if the patient be fed a fat-free diet or is starved, also since foreign 
fats of the food can be identified in the urine; and, finally, since the cells 
in the urine are lymphocytes. 

Lipuria. — Lipuria differs from chyluria chiefly since the urine is opales- 
cent rather than definitely milky. Lipuria is often mentioned in the urine 
reports in hospitals, but by beginners who have not yet learned to exclude 
the oil in the catheterized specimens of urine. The gross or microscopical 
appearances, however, are not sufficient for the recognition of fat. The 
urine should be extracted with ether and the residue examined chemically. 
Fat when heated gives off the odor of acrolein ; the residue will make a fat- 
spot on paper, and will give the osmic acid test. 

The student should exclude deception, the fat used during catheteriza- 
tion, that from the rectum, the tenacious phosphate sediment and the scum 
of bacteria which forms at the surface of the urine. Under normal condi- 
tions there is little if any fat in the urine. Lipuria is said to result from the 
over -ingestion of fat as a food or as a medicine (e.g., cod-liver oil), the so- 
called " alimentary lipuria"; from the subcutaneous injection of oil, or 
an excess of oil rubbed into the skin. Lipuria is said to follow crushing or 
tearing of the subcutaneous fat, injury to the liver, 'or to fatty tumors, 
fracture of the bones, especially if the marrow be crushed and, rarely, acute 
osteomyelitis. It is said to develop in eclampsia, which disease formerly 
was supposed to be due to the crushing of the fat in the pelvis of the kidney. 
Lipuria has been reported in diabetes mellitus, alcoholism, tuberculosis, 



THE URINE: ORGANIZED SEDIMENTS 261 

adiposity, nephritis, certain mental diseases, pancreatic diseases, cardiac 
diseases and after various protoplasmic poisons. A lipemia has been proved 
coincident with fractured bones, subcutaneous bruises, and in diabetes 
mellitus. The claims for other diseases should be confirmed. The slight 
lipuria seen in nephritis in various infections, intoxications, anemias and 
cachexias may be due to fatty degeneration of the kidneys. The fat may 
arise also from epithelial cells, leucocytes, casts and fragments of tumors 
which have undergone fatty degeneration, but under these conditions the 
most of the fat remains in the ceils or collects in droplets which float on 
the surface. 

ORGANIZED SEDIMENTS 

Mucous Sediment. — The ' ' nubecula " is a very faint cloud of mucous 
strands which appear in the top layers of the urine soon after it cools and 
later sinks to the bottom of the glass. This mucus is from the epithelial 
cells of the urinary passages. The mucous strands enclose a few " mucous 
corpuscles," i.e., epithelial cells and mononuclear or polymorphonuclear 
leucocytes, some ameboid in the fresh urine, and some crystals. If 
because of " catarrh " of the urinary passages a good deal of mucus is 
present it may form a definite translucent or cloudy coagulum-like sedi- 
ment better seen after the addition of a little acetic acid. 

Epithelial Cells. — Renal Epithelial Cells. — The epithelial cells 
from the kidney tubules (see Fig. 52, e) are round or cubical in shape and 
larger than leucocytes (12 to 25/x) from which they are easily distinguished 
by their large vesicular nucleus. Their protoplasm is nearly always fatty, 
either finely so or so very fatty that they may resemble colostrum corpuscles 
(c. h.). These cells sometimes show a definite myelin degeneration similar 
to that of the alveolar epithelium cells of the sputum (see Fig. 52, d). 

It is claimed that renal epithelial cells are found in normal urine but 
the chances are that the majority of cells thus reported are endothelial 
leucocytes. Renal cells appear in the urine in all forms of nephritis, but 
especially in the subacute parenchymatous variety in which disease one 
finds them single, in clumps, or attached to casts. In cases of renal infec- 
tion they may be found in the masses of pus-cells (see Fig. 52). 

Hemosiderin in the Urine. 153 — To demonstrate hemosiderin in the urine a fresh 
specimen preferably warm from the body, is centrifugated at high speed and the super- 
natant fluid poured off as completely as possible. The sediment is suspended in the 
trace of fluid that remains and slide preparations studied microscopically for suggestive 
orange or brown granules, more particularly in the cells. A mechanical stage should 
be used and 10 minutes at least given to the search. As a rule the sediment from a 20 c.c. 
specimen will yield a fair number of cells from the higher portions of the urinary tract, 
out often that from 60 to 100 c.c. must be obtained. In urine allowed to cool prior to 
centrifugation the nubecula may prevent proper concentration of the formed elements. 
Kept urines which have become cloudy with urates may be cleared by warming. To 
search a heavy crystalline sediment, or one poor in cells, or containing only leucocytes 

153 Rous, Jour, of Exp. Med., 1918, vol. 28, p. 645. 



262 



CLINICAL DIAGNOSIS 



and squamous epithelium is time wasted. Female patients should be catheterized since 
hemosiderin may come into the urine from the genital tract. 

For the Nishimura test the fresh sediment, as free as possible from urine, is mixed 
with a little human serum untinted with hemoglobin and thick films are made arid dried. 
These are fixed by heat, placed in strong ammonium sulphide for i hour, washed briefly 
with water and subjected for 20 minutes to a fresh mixture of 2% potassium ferrocyanide 
and 1% hydrochloric acid in equal parts. After another brief rinsing with water the 
preparations are stained in lithium carmine for a few minutes, differentiated in acid 
alcohol (1% HC1), and run rapidly through 95% alcohol, absolute alcohol, xylol and 
mounted in balsam. The acid alcohol differentiates the red of the carmine and turns 
the iron granules a deep blue. Its action should be carefully followed with the micro- 
scope since if prolonged it will dissolve the iron, or at least cause the blue tint of the 
latter to run and fade. 

By this method permanent mounts are obtained to be looked over at leisure. The 
iron granules stand out in deep blue against the general carmine tint. 

The presence of cells in the urine containing hemosiderin (the free granules are to 
be disregarded since they may have a different origin) indicates merely the presence of 

an actual siderosis of the kidney paren- 
chyma . Their presence would be important 
in the diagnosis of hemachromatosis in 
cases of doubtful skin pigmentation. These 
cells are present in hemolytic jaundice 
after a siderosis of the kidney has devel- 
oped while their presence would in a doubt- 
ful case speak in favor of pernicious anemia. 

Epithelial Cells from the 
Urinary Passages (see Fig. 51, 6, 
c, d, and Fig. 50, b,f). — A few sur- 
face cells of the transitional epithelium 
of the urinary passages are present 
in the nubecula of normal urines. 
These are increased in number and in variety in inflammatory and destruc- 
tive lesions of this mucosa by the appearance of cells from the middle and 
lower layers of this epithelium. The flat, polygonal, squamous epithelial 
cells (see Fig. 51, e) from the prepuce, end of the ureter, vagina or fossa 
navicularis cannot always be distinguished from the superficial cells from 
the bladder, although usually the stratified grouping of those from the 
vagina makes their recognition easy. 

The cylindrical cells (see Fig. 50, o, e, d) of the urethra are long, narrow 
and bluntly pointed. They occur in pairs or clusters. 

The cells from the transitional epithelium of the urinary passages differ 
in appearance according co the layer from which they originate. Some are 
large and irregular, others round or polygonal. The former are flat, with 
clear protoplasm and usually with a small, very distinct central nucleus. 
Their edges are sometimes very refractile, thin and horny. These are the 
typical pavement cells from the superficial layers of transitional epithelium. 
They are found in large numbers in the urine of patients who are irrigating 
their bladders with too strong fluids, in which cases they may be desquam- 




e, d, cells from male urethra; b, f, cells 
from transitional epithelium; c, shadows of red 
blood-cells. X 400. 



THE URINE: ORGANIZED SEDIMENTS 



263 



ated in large sheets. Dawson 154 who studied such a case found that they 
varied greatly in size and shape. Some were irregular, large and polygonal, 
some smaller and hexagonal. The larger often had a peripheral non- 
granular zone. The nuclei were round or oval, sharply defined and central 
and in many cells were budding. Among these cells were large giant-cells 
with even 1 5 nuclei. In no cells did he see the cupping of the under surface 
so often described. 

The smaller polygonal or elliptical cells from the deeper layers of the 
mucosa have a very granular protoplasm and a large nucleus (Fig. 51, a, 
b, d). Others have a cell body which is definitely oval, or conical, or even 
threadlike, some with 2, 3 or more branches and a very distinct nucleus. 




Fig. 51. — Various forms of epithelium cells in the urine : a, "tailed" cells; b, d, small polygonal; c, large 
surface cells; to the right of d is a small round cell of uncertain origin; e, squamous cells from vagina. 
All of these cells except e were obtained by ureteral catheterization, hence from the pelvis of the kidney 
or scraped from the mucosa of the ureter. The latter is especially true of b, c, d, and neighboring cells, 
which are the forms one gets from normal cases; a, were from cases of pyelitis. X 400. 

These, described as spindle cells or " tailed " cells, were believed formerly 
to come from the pelvis of the kidney. Finally, some from the deeper 
layers of the mucosa are small, round, with a round nucleus and resemble 
mononuclear leucocytes (see Fig. 5 1), which, indeed, some of them may be. 
The best urine in which to study these epithelial cells is that obtained by 
ureteral catheterization. Such cells may arise anywhere along the transi- 
tional epithelium from the pelvis of the kidney to the bladder, singly or 
in clusters. 

The claim that one can tell from the appearance of single cells where 
in the urinary tract they came from is easily disproved. Classes in clinical 
diagnosis should study macerated specimens of this transitional epithelium 



154 Johns Hopkins Hosp. Bull., July 



P- 155- 



264 



CLINICAL DIAGNOSIS 



scraped at various points along the urinary tract. Sahli considered that 
a predominance of tailed cells over other epithelial cells would suggest a 
pyelitis. We agree with this. 

In a case of streptococcus pyelitis the urine obtained at autopsy from the pelvis 
of the kidney contained great numbers of small round polygonal and tailed epithelial 
cells in groups, scores in each field (of 400 magnification) and 3 to 4 of the large polygonal 
cells in each field. Pus-cells were present in great numbers; very little mucus was seen. 

The smaller polygonal cells in groups (Fig. 51, b, d) predominate in 
urine obtained through ureteral catheters. 



% BecKe 



a i 





Fig. 52. — a, pseudo pus-cast; b, epithelial cast showing protoplasmic bridges 

between cells; c, two very granular (myelin ?) renal cells; d, myelin globules; 

e, renal epithelial cells; /, crenated red blood-cells; g, pus-ceLs; h, very fatty 

renal epithelial cells. X 400. 

Casts. — Casts are cylindrical masses of coagulated hyaline or granular 
material moulded into a fairly solid mass, formed in the lumen of the tubules 
and washed out by the urine. They have been classified as: cellular, 
granular and amorphous, but the most are combinations of these. 

Epithelial Casts (Figs. 52 and 55). — The epithelial casts are made up 
in part at least of renal epithelial cells. Some are aggregations of desqua- 
mated cells massed together, with one cell at least well enough preserved 
to be recognized as a renal epithelial cell ; others are actually fragments 
of tubules, with lumen preserved and the cells so intact that even the 
intercellular protoplasmic bridges are visible (Fig. 52). These are the so- 
called " epithelial tubes." In sections of the renal cortex invaginated 
portions of tubules can be seen which, broken off, would be just such casts. 
The cells of epithelial casts may be well preserved, or present all grades 



THE URINE: ORGANIZED SEDIMENTS 



265 



of fatty or granular degeneration. All transitions between these and 
the coarsely granular and fatty casts are seen and, indeed, the most of 
these would be called epithelial could one cell be recognized. If a cast 
which is definitely of some other type, as hyaline, has even one renal 
epithelial cell attached it would be an epithelial cast since it would have 
the significance of one. 

Granular Casts (Fig. 53). — The granules of granular casts may be 
coarse or fine. Of the fine we shall speak later. Coarsely granular casts 
may have the same size and appearance as epithelial casts except no cell 
is well enough preserved to allow its recognition. Some may originally 
have been pus casts. Others are cylinders composed of coarse granules 
with nothing in their appearance suggesting cells. The coarsely granular 
casts have a dense, opaque appearance and dark yellowish color. The 





Fig. S3- — Coarsely and finely granular 
casts. X 400. 



Fig. 54. — Waxy casts. X 400. 



most of the granules are soluble in acetic acid but usually a few fat granules 
are present. The most of these casts probably are made up of the detritus 
of epithelial (or pus) cells which underwent disintegration before or during 
the formation of the cast. (We have no direct proof of the transformation 
of one type of cast to another after the cast has left the kidney.) Hemo- 
globin casts which consist of masses of brownish-red pigment are often 
grouped as coarsely granular casts. 

The finely granular casts present a very different problem. They 
are rather pale and translucent in appearance and seem composed of a 
finely granular material. None of these fine granules are of fat. These 
seem more closely related to the hyaline than to the coarsely granular 
casts and some are hyalines with a few fine granules attached. It would 
seem that the formation of these casts does not necessarily involve the 
actual destruction of entire renal cells, as would seem to be the case of 
epithelial and coarsely granular casts, but rather of their edges lining the 
lumen of the tubules. 



266 



CLINICAL DIAGNOSIS 



Fatty Casts (see Fig. 55).— These striking objects are cylinders made 
up of fatty globules often in clusters which suggest the outlines of the 
original epithelial cells. The granules are yellowish or even blackish in 
appearance and are soluble in ether. Fatty acid crystals project from some. 

Waxy Casts (see Fig. 54). — These casts are composed of a very refrac- 
tive, clear, homogeneous material suggesting wax. They have sharp con- 
tours, are often of a white or yellowish color and show a great tendency to 
split transversely as though very brittle. These may be the longest of all 
casts and extend across the field with twists resembling a corkscrew, or 
the shortest and these suggest fragments of longer casts. Some give the 









Fig. 55. — To the left an epithelial cast with very fatty cells; in the 
center a fatty cast; to the right two leucocyte casts. X 400. 



amyloid reaction, others do not. They are not characteristic of amyloid 
degeneration of the kidney as was formerly supposed and yet in the urine 
of a recent case of amyloid disease it happened that practically every cast 
seen was a waxy cast. In general there are 2 very distinct forms of these 
casts — the yellowish and the bluish. The former, which resemble beeswax, 
were formerly called fibrin casts ; the latter resemble paraffin. Waxy casts 
may be found in the urine of any case of nephritis in which granular casts 
occur and are especially numerous in the scanty urine passed while water 
retention is marked and especially just before death. 

In urine secreted just before death one may see most beautiful waxy casts. On one 
such case the granular and waxy casts were numerous but no hyalines were seen. These 
casts were enormous, many granulars measuring 0.136 mm. and the waxy 0.102 mm. in 
diameter. The latter looked as if made of paraffin. In another case, however, no waxy 
casts were found, only hyalines; yet these were not typical, since too refractile, yet they 
were not waxy. In other such cases all forms were found. 



THE URINE: ORGANIZED SEDIMENTS 



267 




-Hyaline casts of urine. 
X 400. 



In addition to the definitely waxy and hyaline casts are many which 

have not quite the refractility of the former nor do they give the same color 

tests, and yet are more refractile than the hyalines with which they are 

classified. One finds many in some ante- 
mortem specimens. It is hard not to believe 

that they are transitional forms. 

Hyaline, Colloid or Glassy Casts (see 

Fig. 56). — The most common form of casts, 

the hyalines, are so colorless, so translucent 

that they are easily overlooked unless the 

light is almost shut off, or unless crystals or 

cells are attached to them. For this reason 

some as a routine stain urinary sediments 

with Lugol's solution or with analine violet. 

Their outline is very regular, they give the 

microchemical tests for albumin and are 

soluble in acetic acid. They may have the 

same cells attached to them as have all the above-mentioned casts in 

which case the cells give them their name. 

Blood-casts (see Fig. 57). — Blood-casts are either true coagula of red 

blood-cells which have formed within the tubules, or one of the above- 
described casts with at least 1 red 
blood-cell attached. Some of the 
blood-cells are so pale that it is hard 
to recognize them. 

Hemoglobin Casts. — Casts com- 
posed of amorphous masses of hemo- 
globin are seen in hemoglobinuria. 
Others seem to be hyalines or granular 
casts impregnated with hemoglobin. 
Pus-casts (see Fig. 55) are similar 
in the method of their formation to 
the blood-casts. The pus-cells appear 
more spherical than the epithelial but 
to be certain of their nature one 
should make their nuclei evident by 
adding acetic acid. The most of the 
pus-casts are hyaline or granular casts 
with at least 1 leucocyte attached. 

Those which are conglomerates of pus-cells are rare. 

Cylindroids (see Fig. 58). — The threads of mucus in the urine are 

either the so-called " mucous threads," which are flat ribbons of mucus 

which do not at all resemble hyaline casts, which often extend over several 

fields and vary much in diameter along their course (such threads make 




Fig. 57. — Blood-cast. X 400. 



268 CLINICAL DIAGNOSIS 

up the nubecula) and the so-called cylindroids which may closely resemble 
casts. These differ in appearance from hyalines in that one or both ends 
taper off into a longer or shorter thread. These have not the fibrillar 
appearance of mucous threads and chemically they resemble casts. Since 
they occur where tine casts would be expected some claim that they have 
the same significance as they. The problem is a difficult one but we call 
no structure a true cast until we are sure that neither end runs off into a 
thread. Cylindroids covered by urates have the appearance of granular 
casts. The point is an important one, for if they are mucous threads 
they certainly arise from the mucous surface, while if casts they should 
arise in the renal parenchyma. True mucous threads arise in the blad- 




FlG. 58. — a, cylindroids, i.e., bodies much resembling hyaline casts; b, mucous 

threads; c, a spiral structure of material resembling hyaline casts or mucous threads; 

d, a vegetable thread. X 400. 

der chiefly. One seldom sees them in urine catheterized from the pelvis 
of the kidney. They are insoluble in acetic acid while many cylindroids 
are soluble. 

Combined Casts. — A cast may be clear, therefore waxy or hyaline, 
at one end and granular at the other; or it may have cellular elements 
attached. All combined casts take their name from the cells attached and 
if a variety of cells are involved, from that one which would be most 
important in prognosis. 

Bacterial Casts. — Bacterial casts are masses of bacteria in the shape 
of a cast which would seem to be moulds of the tubules. These occur in 
purulent infectious pyelonephritis and in pyemic kidneys. Other casts 
may become permeated by bacteria in a remarkably short time. 

Urate Casts. — In the urine of the new-born with uric acid infarcts 
of the kidney may be found casts which are masses of sodium urate. 



THE URINE: ORGANIZED SEDIMENTS 269 

Pseudo-casts made up of urates are common. Other casts in a con- 
centrated urine may become incrusted with urates and hence be more 
dark, homogeneous and granular then true granular casts. The urate 
masses also have uneven edges and disappear on warming. Scratches in 
the glass are sometimes confusing (Fig. 59). In some cases of pyuria 
(cystitis, e.g.) the mucus threads full of pus-cells make very perfect pseudo- 
casts (Fig. 52). 

The length of true casts varies from very small fragments to 1 mm. or 
longer. Some are narrow, others broad. From the size of the casts no 
conclusions can be drawn of their source so much does the size of the 
tubules vary in pathological conditions. Some can almost be seen with 
unaided eye. It was formerly supposed that the beautiful corkscrew forms 
so often seen come from the convoluted tubules, but this is improbable 
since any corkscrew shape would probably be 
effaced during their passage through the straight 
tubules. Some are spiral all their length, others 
only at one end. This, says Senator, merely 
shows that they are composed of a tough elastic 
material and have been squeezed through a nar- 
row orifice. The end of the cast is seldom 
split or forked. 

The origin of epithelial casts, especially of 
those with a lumen, is not disputed; nor is that 
of those blood- and pus-casts which are con- 
glomerates of cells. The coarser granular casts 
quite certainly are made up from the detritus of 
epithelial or pus -cells. All transitions from the fi G - 59.— Pseudo-casts. From 

left to right, a linen thread, a 

coarsely granular to the waxy casts may be vegetable spine, a cotton thread, 

■ . . . . and a scratch on the glass slide. 

found, especially in sections of kidneys. The 

origin of the hyaline casts, however, has long been in dispute. Some claim 
that they are a coagulated exudate from the blood into the tubules, others 
a product of the secretion of the epithelial cells themselves; that is, the 
slightly injured epithelial cell may furnish an abnormal secretion of coagula- 
ble material which coagulates in the tubules. Hyaline globules can be 
demonstrated in these cells and the confluence of these in the tubules would 
explain casts. This is the generally accepted idea. Another explanation, 
for which the study of microscopic sections of kidneys gives evidence, is 
that the fine edge of these epithelial cells undergoes hyaline degeneration, 
sloughs off and is moulded into a cast. Hyaline casts are certainly not 
composed of coagulated fibrin. (They arise where there is no suspicion of 
inflammation, as in a practically normal kidney, e.g., the albuminuria of 
the new-born; they do not give the Weigert's fibrin stain.) They are not 
simply coagulable albumin, for where this is present, as in cases of chyluria, 
these casts may be absent. There is no evidence that the albirmin of the 




270 CLINICAL DIAGNOSIS 

blood is their source, since albumin and casts are little if at all related in 
their origin, for either one may be present without the other. There is 
evidence that if hyaline casts remain unusually long in the tubules they 
become waxy (see page 266). 

The origin of hemoglobin casts is an interesting problem since hemo- 
globin is soluble in urine. It may be, however, that these are hyaline or 
waxy casts impregnated with hemoglobin. 

The chemistry of casts has been but little studied. Probably none are 
composed of fibrin. Whether any consist of amyloid or not is disputed. 
Very few give a typical amyloid reaction, the majority of those which look 
waxy taking merely a brownish color with Lugol's solution and a reddish 
color with gentian violet. Most genuine casts are soluble in acetic acid, 
and this is a valuable aid in distinguishing them from mucous cylindroids. 

Diagnostic Importance of Casts. — It is claimed that casts sometimes 
appear in the urine of persons whose kidneys are quite normal. This is 
not generally admitted. The disturbance leading to the cylindruria may 
be but slight and transitory but their presence is always evidence of an 
abnormal condition of the renal epithelium. This may be a temporary 
disturbance of the renal circulation, a temporary condition of malnutrition, 
a mild irritation, congestion, pressure, etc. ; or the cause may be permanent 
and serious, as in nephritis. It is quite true that from the number and 
character of the casts present -in the urine one cannot judge as to the 
severity of the cause of the cylindruria. Indeed, it would seem that, the 
more normal the condition of the cells prior to the renal disturbance the 
more brilliant will be the display of casts. Certainly the most seriously 
injured (small, contracted) kidneys may for fairly long periods excrete 
urine containing few or none. 

The general statement may be made that casts appear under the same 
circumstances and have the same general significance as albumin, although 
it would seem that of the two the albumin is a little more sensitive index 
of renal disturbance than are casts. 

While albuminuria and cylindruria are usually associated, either may 
appear without the other. Casts have been reported absent when albumin 
was present in some cases of chronic nephritis (especially the arteriosclerotic 
type), in some cases of jaundice and in some of febrile albuminuria. To 
prove casts absent, however, is no easy matter since in some urines they 
disintegrate very rapidly. The search should be made with the aid of a 
centrifuge while the urine is very fresh. The rapid disappearance of casts 
has been ascribed to the presence in the urine of a ferment which some claim 
is pepsin, others, a bacterial enzyme. 

On the other hand, it is not at all rare to find casts in urines in which 
albumin cannot be demonstrated by clinical methods. A pure cylindruria 
may be due to som.e food, as asparagus, radishes, coffee and mustard; or 
to a drug, as alcohol, salicylic acid, mercury, arsenic and camphor; or to 



THE URINE: ORGANIZED SEDIMENTS 271 

an injection of tuberculin, etc. Pure cylindruria is sometimes seen in cases 
of heart disease, cancer, jaundice and constipation; in acute infectious 
diseases, as scarlet fever, typhoid fever, erysipelas and tuberculosis; in 
nephritis, especially chronic, in acute cases especially during convalescence 
and even in uremia. In fact, in any case in which albuminuria would be 
expected cylindruria alone may occur. It is of interest that among the 
athletes studied with reference to " physiological albuminuria " were 
several with pure cylindruria. In a recent case of brain tumor we found 
in the urine many epithelial and waxy casts and many cylindroids, but 
no albumin. 

In many cases there is little doubt but that both casts and albumin 
were for a time present, but that the latter disappeared first. We believe 
that the casts are apt to disappear before the albumin in those cases in 
which the renal lesion is chiefly degenerative, as in patients with arterio- 
sclerosis; the albumin first, in those cases in which the lesion is inflamma- 
tory, as in parenchymatous nephritis. 155 Without much doubt our failure 
to find more cases of pure cylindruria has been due to our habit of not 
examining the sediment carefully if the test for albumin is negative. 

The association of casts and " nucleo-albumin " but no serum albumin 
is often noted (see page 222). 

In any given case of nephritis there usually is a rough parallelism 
between the number of casts and the amount of albumin in the urine, but 
the casts are a much more variable factor than is the albumin. They are 
most abundant in acute and subacute parenchymatous nephritis, and 
fewest in interstitial nephritis, amyloid disease and chronic passive con- 
gestion. There is no evidence as yet for a special form of nephritis with 
cylindruria its most prominent urine feature. In the diagnosis of the 
grade of a case of renal trouble the number and variety of casts are not as 
important as are the specific gravity of the urine, its chemical analysis, 
the blood chemistry and especially the history and physical condition of 
the patient. Casts, together with albumin, would seem to depend more on 
the acute element of the process called nephritis than on the total lesion. 
There are no casts which are pathognomonic of nephritis ; any or all varieties 
may appear in the urine of patients with nephritis and also of patients 
who have no true renal disease. But nephritis is practically the only 
condition in which cylindruria is long-continued; in other cases it lasts 
but a few hours or days. 

" Showers " of casts, i.e., the sudden appearance of numbers of casts 
greatly in excess of that of the preceding or of the succeeding days, usualfy 
last but part of a day. These may occur in any form of nephritis. 

In the cylindruria not due to nephritis the casts are as a rule few and 
these usually are hyaline. But in the urine of athletes following great 

155 Emerson, Jour, of A. M. A., Jan. 6 and 13, 1906; Vincent, N. Y. Med. Jour., 
April 13, 1907. 



272 CLINICAL DIAGNOSIS 

strain all varieties may be found (see page 225). Epithelial and leucocyte 
casts are not nearly so rare as is imagined and may occur in even non- 
inflammatory transitory cylindrurias. 

It was formerly supposed that the presence of epithelial and the hyaline 
casts meant an acute process, that of granular and waxy casts a more 
chronic process ; but in all forms of nephritis all kinds of casts may appear ; 
in amyloid disease even there is nothing characteristic in the urine picture. 
Sahli suggests that casts in the forming become granular and waxy casts 
from lying a long time in the tubules and that this explains the large num- 
bers of these present in the urine after a period of suppression, as after an 
acute nephritis or an acute exacerbation of a chronic form. We have 
noticed this also in other oligurias; that, for instance, following decap- 
sulation of the kidneys. During the first few days after the operation 
as the urine begins to increase in amount a very large number of waxy 
casts appear. 

Almost any kind of casts may appear following various renal disturb- 
ance. Brown 156 has reported some interesting results of operation on 
normal kidneys (nephropexy or exploratory nephrotomy). Cn the first 
day after the operation the urine contained casts in enormous numbers, 
hyaline, granular and epithelial. Considerable albumin also was present. 
These casts rapidly diminished in number and in from 2 to 6 days entirely 
disappeared. During this time there were no symptoms of nephritis, no 
edema and no change in the amount of urine. The disproportion between 
the small amount of albumin and the great number of casts was a marked 
feature of these casts. There were no later symptoms. 

Casts in great numbers are important as a prodromal symptom of dia- 
betic coma (Kulz). These may appear in immense numbers before the 
coma begins and even form a macroscopic sediment. These casts are 
characteristic in appearance — short, broad, of delicate outline, pale, the 
most of them granular and hyaline and with few other formed elements. 157 

Staining Casts. — All methods of staining casts are unsatisfactory, 
because the stain precipitates in the urine or the albumin of the urine may 
itself take the stain. The specimens cannot be dried for this reason. The 
best method is to wash the casts 1 or 2 times by sedimentation with 0.6% 
sodium chloride solution to rid them of all soluble matter and albumin. 
In the next centrifugalization 1% methylene blue may be added. To 
hasten centrifugalization a little alcohol should be added, not much, nor 
should it be allowed to remain for ? long time in contact with the sediment 
else a coagulum will result. 

To preserve the casts and also to stain them, they should be washed as 
above in normal salt solution and lastly in a 1% osmic acid solution, or 

156 Johns Hopkins Hosp. Bull., May, 1900. 

157 See Domansky and Reimann, Zeitschr. f. Heilk., 1901, and Herrick, Am. Jour. 
Med. Sci., vol. cxx, 1900. 



THE URINE: ORGANIZED SEDIMENTS 273 

in i to 10% formalin, or in a 5% HgCl 2 solution for 5 minutes. In the latter 
case they are then washed with water and preserved in from 2 to 10% (or 
1 to 2%) formalin. If no red blood-cells are present the mercuric chloride 
should not be used since it disturbs microchemical tests. In case formalin 
is used the casts should be especially well washed or the spherical crystalline 
masses of diformaldehydurea will form. Gumprecht adds that it is not 
really necessary to wash the casts if they are well centrifugalized and the 
supernatant fluid completely decanted. A good staining method for fat 
and cell nuclei was described by Cohn. 158 The specimen, well washed by 
centrifugalization in normal salt solution, is air-dried on the cover-glass 
and hardened by immersing the glass in 10% formalin for 10 minutes. It 
is then washed rapidly but gently with H 2 and then immersed for 10 
minutes in a concentrated Sudan III solution in 70% alcohol. It is then 
washed in 70% alcohol for 1 to 2 minutes and then stained briefly in hema- 
toxylin (Ehrlich's solution). The specimens are mounted in glycerin. 

Koslowoski 159 recommended Farrant's mounting fluid. This consists 
of equal parts of water, glycerin and saturated aqueous solution of arsenous 
acid (saturated by weeks of standing); to this gum arabic, }{ volume, is 
added and this allowed to stand (about three weeks) till all is dissolved. It 
is then filtered if necessary. The urine is mixed in a centrifuge tube with 

1 cm. of t% eosin or methyl violet, then centrifugalized and washed by 
centrifugalization till all the urine is removed. One drop of the sediment 
is then mounted on the slide with 1 drop of the above fixing fluid. 

Bohland advises to wash the sediments with salt solution and then to 
add Miiller's fluid. They are kept in this for 2 weeks, changing the fluid 

2 or 3 times. The Miiller's fluid is then decanted and the sediment washed 
in absolute alcohol until this is colorless. 

Testicular Casts. — Casts have been described in cases of " spermator- 
rhea " which " can hardly be distinguished from renal casts except that the 
urine is otherwise normal. They are all in the first glass of the two-glass 
test and the presence in the same specimen of spermatozoa will indicate 
their origin. They are supposed to arise in the testicle. ' ' We have inquired 
of those with a very wide experience in the examination of prostatic secre- 
tions and they say they have never seen any such objects, although certain 
cylindrical cells may at first glance resemble true casts (see Fig. 64). 
Spermatozoa may often be found, active at first, together with all of 
the elements of unripe semen. They soon disintegrate. Such are found 
not only after coitus and pollution, but also after epileptic and other 
convulsive seizures. 

Gonorrheal Threads, Clap Threads. — These threads occur in a late 
stage of acute gonorrhea and in chronic gleet after the exudate becomes 
very mucous and scanty and so collects in the longitudinal furrows of the 

158 Zeitschr. f. klin. Med.,- 1899, Bd. 38. 

159 Virchow's Arch., 1902, vol. clxxix, p. 161. 

18 



274 CLINICAL DIAGNOSIS 

mucosa. They may be from a few millimeters to one centimeter long and 
are yellow or white in color. Some, and these are found in very chronic 
cases, are narrow, delicate, transparent threads which consist of mucus 
in which are imbedded a few epithelial and still fewer pus-cells. Others 
are shorter, firmer and contain more cells, especially pus-cells. They 
settle at once to the bottom of the glass, but float up as fine, easily recog- 
nizable threads if the urine be agitated. They may coalesce in the cpurse 
of time and so become unrecognizable. 

Tissue Fragments. — Portions of carcinoma have been found in the 
urine, especially from papillomatous cancers of the bladder, some of which 
were large enough to be sectioned for microscopic study. In some of these 
spindle-cells were found which enclosed hematoidin crystals and red blood- 
corpuscles. Fragments from renal cancers have been very rarely found. 
No mass of recognizable sarcoma tissue has as yet been recovered from the 
urine, but Rothschild 160 found a structureless mass in the urine of a case 
of giant-cell sarcoma of the kidney which was 5.2 cm. long, 0.5 cm. wide, 
firm, glossy and transparent. Masses of caseous matter are sometimes 
found in the urine in cases of renal tuberculosis. To demonstrate elastic 
tissue fibers the urine should be centrif ugalized, acid added to dissolve the 
phosphates, the supernatant fluid decanted and the sediment then warmed 
with an equal amount of 10% KOH, which will destroy all but the elastic 
tissue. The specimen is then again centrif ugalized and the sediment 
examined microscopically. 

Other gross masses which are met with in the urine are mucous casts 
(see page 221) and the fibrin masses sometimes found in chyluria, hematuria 
and in inflammatory conditions, especially tuberculosis. 

Pus-cells. — A few pus -cells might perhaps be expected in any normal 
urine, but the presence of many would mean inflammation of the urinary 
passages, of the kidney, or the rupture of an abscess into the urinary tract. 
The number of pus-cells varies enormously. As a rule, if from the cortex 
of the kidney they are few in number ; if from the passages, many. 

Hottinger found in a case of cystitis 150,000 leucocytes per cubic 
centimeter of urine, which would be a daily loss of about one per cent, of 
the total number of leucocytes present at any one time in the normal 
circulating blood. 

The origin of the pus-cells in a specimen of urine may be indicated by 
other constituents present, i.e., by the character of the epithelial cells also 
found, by the casts, etc. The sudden appearance of a large amount of 
pus usually means a ruptured abscess. The pus-cells in gonorrhea are often 
enclosed in threads of mucus, the so-called Tripperfaden (see page 273). 

The urine of women usually contains pus from the vagina. 

The pus in alkaline urine swells to a slimy, gelatinous mass, which is 
more slimy still since it usually is mixed with so much mucus. Microscopic- 

160 Deutsches med. Wochenschr., 1901, No. 50. 



THE URINE: ORGANIZED SEDIMENTS 275 

ally, the pus-cells in very acid urine are so cloudy and shrunken that they 
are unrecognizable unless acetic acid is added to make their nuclei visible. 
In an alkaline urine they swell and become glassy but even then it is not 
easy to see their nuclei. In a weakly alkaline, amphoteric, or faintly acid 
urine they may remain well preserved for a long time and even show active 
ameboid motion. The diameter of leucocytes varies from 7 to 12/x and 
their nucleus is small, usually polymorphous. The nuclei of some are 
vesicular. It is hard to distinguish these last cells from renal epithelial 
cells unless the sediment be stained. Senator considered that many of the 
pus-cells in Bright 's disease were mononuclear leucocytes. 

In the urine sediments of 2 patients all the leucocytes were so drawn 
out that they resembled spindle epithelial cells. This probably was due 
to too long centrifugalization. It often, but that was not true of these 
cases, is due to the technic of making the slide specimen. 

The Albumin of a Purulent Secretion. — There will always be 
some albumin in solution in a urine with a pus sediment (and this cannot 
be removed by nitration). It often is important to decide whether more 
albumin is present than the pus serum can explain; that is, whether a true 
renal albuminuria also is present. If casts and renal epithelium are found, 
a cortical origin for some at least of the albumin may be assumed. 

Posner's Method. — The total albumin of the filtered urine is first care- 
fully estimated. The urine (a 24-hour specimen) is then well shaken 
and the leucocytes counted, using the leucocyte pipet and Toisson's solu- 
tion and the ordinary blood-counting chamber. For each 100,000 leuco- 
cytes per 2 c.c. of urine, one may expect 0.1% albumin (Goldberg, 2 p.m.). 
If over 3000 cells per cubic millimeter are present the urine should be 
diluted with a 1 to 3% NaCl solution to bring the count near this figure. 
Kretschmer 161 believes this method valuable in following the treatment 
of a case. 

Posner described also an easier method which has some value. The 
well-shaken urine is poured into a flat-bottomed beaker which rests on a 
sheet of ordinary printed paper until the urine so obscures the type that 
it can no longer be read. One can read through a layer of normal urine 
8 cm. deep. If the type ceases to be legible when the urine layer is from 
0.5 to 1 cm. deep we may conclude that 40,000 leucocytes per cubic centi- 
meter are present; if 6 cm., 1000 leucocytes per cubic centimeter. This 
method is of some value in following the success of treatment. 

Donne s Pus Test. — The supernatant urine of a specimen with a sedi- 
ment which suggests pus is poured off and to the sediment is added a small 
piece or a strong solution of KOH or NaOH. If the sediment is pus it 
will be transformed to a viscid gelatinous mass which sticks to the glass. 

Pus-cells will take a mahogany-brown color if treated with Lugol's 
solution. 

161 Jour. A. M. A., 1917, vol. lxix, p. 1505. 



276 CLINICAL DIAGNOSIS 

Red Blood-corpuscles in the Urine in Cases Without True Hematuria.— 

Red blood-cells are present in the urine in cases of acute trauma, of 
stone anywhere along the urinary tract, in chronic passive congestion, in 
the hemorrhagic diathesis, after severe exercise, as long foot-races (Barach) 
and in many trivial conditions in which they would not be expected. 
vSome of the cells are intact, others are shadows. 

In concentrated urines the red blood-cells are crenated; in dilute, they 
are swollen or laked; in acid urines, intact; in alkaline, they are destroyed 
and form masses of yellowish-brown granules. 

It is important to decide whether in any given case the red cells in the 
urine come from the cortex of the kidney or from some point along the 
urinary tract. That they come through the cortex may be assumed if 
many red blood-cells are sticking to casts or if true blood-casts also are 
present. An amount of blood sufficient for large clot formation seldom 
comes through the cortex and yet in rare cases of nephritis and in some 
cases of so-called renal epistaxis the urine may contain large blood- clots 
having the shape of the renal pelvis or of the ureter. 

Gumrecht claimed that if many of the red cells were fragmented, that 
is, present as clumps of granules, one may assume that they were from the 
cortex since urea is the only constituent of the urine which could fragment 
them and the urea solution is strong enough (8%) to do this only in the 
cortex. Goldberg, however, believes that red cells can become fragmented 
in an infected bladder. 

CONCRETIONS 

Renal and Bladder Stones. — By renal stones are meant concretions 
from the pelvis of the kidney and the ureter. These vary in size from a 
grain of sand to large arborescent concretions which fill the whole renal 
pelvis. One weighed 1088 grains. The branches of these large stones may 
be hollow, thus furnishing a passage for the urine. The bladder concretions 
are single or multiple and vary greatly in size. 

Uric Acid Concretions. — Of the renal stones those which are composed 
chiefly of uric acid are the most common. The size of those found in the 
bladder varies from that of a pea to that of a goose egg. They are always 
colored, their tint varying from a grayish-yellow to a yellowish or pale 
reddish -brown. Their surface is sometimes smooth and polished, sometimes 
rough and nodular. They are very hard, fracture easily and on cross-section 
show a crystalline structure and concentric arrangement of layers of differ- 
ent colors, which layers may be composed alternately of uric acid and some 
other salt, as CaOx. These stones burn without residue if pure; they give 
the murexid test ; on the addition of NaOH they liberate but little ammonia ■ 
they are soluble in alkali, and from this solution, if acetic acid be added, 
crystals of uric acid will crystallize out. These crystals should be subjected 
to the murexid test. 



THE URINE: CONCRETIONS 



277 



TABLE IV 

When the Concretion is Heated on the Platinum-foil the Powder 
(hofmeister's table) 



Does not burn 


Burns 


The powder + HCl 


With flame 


Without flame 








CD O 


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Pure ammonium urate stones as primary concretions are found chiefly 
in the new-born, rarel] in adults, although as secondary deposits they are 
very common. These stones are almost as soft as wax and when dry are 
clayey and easily powdered. They give the murexid test and if heated 
with NaOH much ammonia is liberated. 



278 CLINICAL DIAGNOSIS 

Calcium oxalate stones are, next to uric acid stones, the most com- 
mon. Since they are the hardest and heaviest of stones they easily cause 
hemorrhages and so are often stained dark brown with blood pigment. 
They are soluble in HC1 without gas formation, not in acetic acid, but if 
moderately heated the powder produced is soluble in acetic acid with gas 
evolution. If heated to a high temperature the powder reacts alkaline 
because of the.Ca(OH) 2 formed. Kahn 162 who studied 16 renal stones 
decided that the most were composed of calcium oxalate; that they all 
contain some uric acid and urates ; and that the shape, color and consistency 
of a stone give but little evidence of value as to its composition. 

Phosphate Stones. — Pure phosphate stones are found rarely in the 
pelvis of the kidney. Those which are are small and consist of a mixture 
of the normal phosphates of the alkaline earths and the triple phosphate. 
In the bladder these stones may grow to very large size. The phosphates 
are also common ingredients of mixed stones. Phosphate stones often 
form around a foreign body. Their color varied — sometimes they are 
white or pale yellow, or purplish. They are soft, of light weight and have 
always a rough surface. The rare concretions of pure triple phosphates 
are small with a granular surface, upon which are often encrusted red 
crystals. Stones of acid calcium phosphate, which are rare, are white and 
of a beautiful crystalline structure. Phosphate stones if powdered do not 
burn, are soluble in acetic acid without gas formation and in this solution 
can be found phosphoric acid and the alkaline earths. They usually con- 
tain a great many organisms. The triple phosphate stones liberate much 
ammonia on the addition of NaOH. 

Calcium carbonate stones which are rare in man, are chalky white in 
appearance and soluble in acid with gas formation. 

Cystin stones are rare, but 106 cases having been reported. 163 Renal 
cystin concretions are seldom larger than a small pea, but those in the 
bladder may become as large as a hen's egg. They are light in weight, 
smooth and often so soft and wax-like in consistency that they may be 
crushed between the fingers. They have a smooth or ragged surface, are 
white or pale yellow in color, crystalline on cross-section, burn readily and 
perfectly on a platinum-foil with a bluish flame and are soluble in ammonia 
and recrystallized by acetic acid (for the other reactions of cystin, see 
page 256). 

The very rare xanthin stones as a primary formation form occur 
especially in children. They vary in size from a pea to that of a hen's egg. 
They are pale white or yellowish-brown in color, rather hard, amorphous 
on cross section and on rubbing appear like wax. They burn without 
residue on the platinum-foil and the material of which they are composed 
gives the reactions of xanthin. 

162 Arch, of Int. Med., January 15, 1913, vol. xi, p. 92. 

163 Kretschmer, Urolog. and Cutan. Rev., 1916, vol. xx, No. 1. 



THE URINE: BACTERIOLOGY 279 

Fatty Concretions. — Only a few fatty concretions have been reported. 
These contained free fatty acid, neutral fat and much cholesterol. Some 
proved to be composed of the fat used in passing bougies. 

Indigo. — Three stones, composed in part of indigo, are on record and yet indigo 
may be the nucleus of various other stones. They have a blue or bluish-gray surface. 
Albumin. — One calculus said to consist of albumin is on record. 

THE BACTERIOLOGY OF THE URINE 

Unless extraordinary precautions are observed in collecting and pre- 
serving urine it soon contains hosts of bacteria of many varieties. Some of 
these it may have contained when it was voided, but the most are contami- 
nations from the external genitalia, from the vessels which hold the urine 
and from the air. The urine is an excellent culture medium for many 
organisms and they soon render it unsuitable for chemical or microscopical 
study. Specimens to be studied chemically should be collected in clean 
bottles, chloroform, camphor, thymol, formaldehyde, etc., should be added 
at the very first and the specimen kept in an ice chest as much of the time 
as possible. Specimens to be studied microscopically should, whenever 
possible, be examined at once after it is voided. Especially if the specimen 
is to be studied bacteriologically, special technic is necessary in collecting 
and keeping it. 

The Technic of Obtaining Specimens for Bacteriological Study. — To 
obtain a specimen for bacteriological study it is not always or often neces- 
sary to catheterize the male patient, especially if he is intelligent enough 
to observe the necessary precautions. The glans penis, and especially the 
edges of the urethral orifice, should be washed thoroughly with green soap 
and water and then with bichloride of mercury (i : iooo). The anterior 
urethra is then thoroughly irrigated with bichloride of mercury (i : 60,000). 
The patient then voids; the most of the urine is allowed to escape, thus 
completing the irrigation of the tract, and the last few cubic centimeters 
are collected in a sterile test-tube. A way preferred by some is to ask the 
patient to void into three sterile glasses. The third contains the specimen 
to be examined. 

It is always necessary to catheterize the female patients. The external 
genitalia, and especially the orifice of the urethra, are well washed with 
green soap and water. The orifice of the urethra is then repeatedly mopped 
with sterile cotton pledgets soaked in sterile water, boracic acid, or mercuric 
chloride. At least 10 or 12 of these pledgets should be used. A sterile glass 
catheter is then inserted with care that it touches only the orifice of the 
urethra. The hands of the person introducing it should be surgically clean. 
Over the free end of the catheter should be fitted a rubber tube which will 
protect the tip of the catheter from contamination. This should be large 
enough to fit loosely and be about 4 inches long. After the most of the 
urine has escaped, this rubber tube is slipped off and the last small portion 
of urine collected in a sterile test-tube. 



280 CLINICAL DIAGNOSIS 

Bacterioscopic Examination of the Urine. — The examination of a smear 
made immediately after the specimen is obtained is by far the most import- 
ant part of a bacteriological examination of the urine since in this way we 
may get a hint as to what culture media will best serve our purpose and 
may discover bacteria which will not grow on the media used as well as 
those which have already died. The smear may show a rich flora and the 
cultures the reverse. 

Two of the reasons why smears of urinary sediments are not oftener 
studied for bacteria are that it is difficult to obtain good film preparations 
unless all the urea, which is very hygroscopic, has been previously washed 
out from the sediment ; and secondly, that it is difficult to sediment bacteria 
by centrifugalization since the specific gravity of their bodies is nearly the 
same as that of urine. Nevertheless in the great majority of cases, especi- 
ally if there is even a little pus present, one does get good smear prepara- 
tions. One centrifugalizes the urine on a rapid machine until there is even 
a little sediment at the point of the tube, then quickly inverts the tube and 
allows all the urine to escape and drain. While still holding the tube in 
a perfectly vertical position a little of the sediment is scraped from the tip 
of the tube with a platinum loop. One must be careful to invert the tube 
to the vertical position quickly, and while the urine is draining and while 
obtaining the sediment not to incline the tube at all, else urine clinging to 
the sides may flow to the point and so add urinary salts to the smear. 

But in case the urine is very clear and one wishes to obtain on a film 
preparation (but not for cultures) any organisms which may be present, 
one dilutes it with one, or even two, volumes of alcohol. This will so lower 
its specific gravity that practically all the organisms will be thrown by the 
centrifuge to the point of the tube. The urine thus diluted is very thor- 
oughly centrifugalized, the supernatant fluid poured off, more alcohol or 
distilled water added, the contents of the tube well shaken and then again 
centrifugalized. (There is danger that many of the organisms will be left 
sticking to the sides of the tube rather than be thrown to its point. To 
obviate this in part at least, some allow a considerable amount of the mix- 
ture of urine and alcohol to sediment by gravity in a beaker and then centri- 
fugalize the sediment.) The sediment will now be free from urea and satis- 
factory smear preparations on a glass slide or cover-glass can be made. It 
is often wise, in case but little sediment is present, to add a little egg 
albumin to stick the bacteria to the glass. 

The smear preparation is first dried in the air, then passed slowly 
through the flame of a Bunsen burner or alcohol lamp three times and 
then stained. 

Bacterial Stains. — The bacterial stains in common use are solutions 
of the basic aniline dyes. 

Lbfflers Methylene Blue. — Saturated alcoholic solution of methylene 

e 30 c.c. and aqueous solution of KOH (1 : 10,000) 100 c.c. 



THE URINE: BACTERIOLOGY 281 

The film is covered with this stain and heated over the flame for from 
i to 5 minutes. When no heat is used the staining will take much longer. 
The stain is then washed off with water, the film dried with blotting or 
filter paper and then mounted in Canada balsam. If on a slide the smear 
may be studied without the interposition of a cover-glass. 

Saturated Aqueous Solution of Methylene Blue. — This is used as the 
above but stains a little more slowly. 

Aniline Gentian Violet. — Aniline oil water is first made by adding 
exactly 2 c.c. of aniline oil to 98 c.c. of distilled water in a flask. This is 
shaken vigorously till as much as possible of the oil is dissolved and then 
filtered twice through the same paper. This fluid is kept in a dark-glass 
bottle and in a dark place. 

To 75 c.c. of this aniline oil water are added 25 c.c. of a saturated 
alcoholic solution of gentian violet and the mixture filtered. This 
staining mixture is fairly permanent but should not be exposed to 
strong sunlight and should be occasionally filtered. Smears will stain 
readily in this in a few minutes. This is the stain used in Gram's method 
(seepage 38). 

Pijfaud's Method 164 of Staining Bacteria. — This is a valuable method 
for determining the nature of a growth. 

Cyanide blue solution: 

Distilled water 100 parts; 
Potassium cyanide (pure) 1 part; 
Potassium carbonate (dry; pure) 0.5 part; 
Rectified methylene blue 0.5 part. 
A small drop of this stain is placed on the center of a slide, and then a 
loop of the growth well mixed with it. After 1 minute a clean cover-glass 
is dropped on this fluid and the excess of the moisture absorbed by pressing 
the cover-glass firmly with a piece of filter paper. In this way one dis- 
penses with drying, heating and long staining. 

Carbolfuchsin. — This contains : 
Basic fuchsin 1 part; 
Absolute alcohol 10 parts; 
Carbolic acid solution (1 : 20) 100 parts. 

This is a very powerful stain which when undiluted will stain bacteria 
in from % to 1 minute. Better results are obtained if it is diluted with from 
5 to 10 volumes of water and left in contact with the smear for a few 
minutes. This is the stain used for the tubercle bacillus (see page 25). 

Bismarck Brown (see page 38). 

Staining Methods for Acid-fast Bacilli (see page 25). 

Gram's Method (see page 38). 

Capsule Stains (see page 32). 

164 N. Y. Med. Jcur., Nov. 2, 1907. 



282 CLINICAL DIAGNOSIS 

Spore Staining. — The film is first placed in a jar of chloroform for 
2 minutes and then well washed in water. It is next placed in a 5% solu- 
tion of chromic acid for from % to 2 minutes and again well washed with 
water. It is then covered with carbolfuchsin and heated in the same 
manner as if one were staining the tubercle bacillus (seepage 25). The 
carbolfuchsin is not washed off with water, but with 1% sulphuric acid, or 
with methylated spirit (ethyl alcohol 9 parts, methyl alcohol 1 part) and 
left in this until decolorized. It is then washed in water, stained with a 
saturated aqueous solution of methylene blue for % minute, washed again 
in water, dried and mounted in balsam. The spores will retain the red 
stain, the bacilli will stain blue. 

Flagellum Staining. — It is very difficult to get good specimens of 
stained flagella since only under certain conditions of growth, age, etc., 
can the flagella be demonstrated and even when the culture is a suitable 
one the flagella may very easily be injured by the technic used. The smear 
to be studied should always be made from a young agar culture incubated 
at 37 C. for from 12 to 18 hours. Kendall recommends to inoculate gently 
5 c.c. of sterile water with enough of the above-mentioned growth to produce 
a faint turbidity in the upper half of the tube. This tube is then placed 
in a thermostat for 1 hour. This will allow the clumps to settle and the 
organisms to multiply a little. Without disturbing the fluid any more 
than one can help 2 or 3 loopfuls are placed on a clean cover-glass without 
attempting to spread the fluid at all and dried in a thermostat. The speci- 
men is then fixed in a flame. (The cover-glass should be one which had 
been washed in a mixture of concentrated sulphuric acid 6 parts, potassium 
bichromate 6 parts and water 100 parts. It should then be washed thor- 
oughly in water and kept until used in absolute alcohol.) 

The staining methods are all of them so unsatisfactory that the best is 
usually the one with which the worker is most familiar. 

Pitsfield's Method as Modified by Richard Muir. — The mordant con- 
sists of: 

Tannic acid, 10% aqueous solution, filtered, 10 c.c; 
Alum, saturated aqueous solution, 5 c.c; 
Corrosive sublimate, saturated aqueous solution, 5 c.c; 
Carbolfuchsin stain (see page 281) 5 c.c. 
This is mixed thoroughly. A precipitate forms which is allowed to settle, 
or the fluid is centrifugalized, and the clear supernatant fluid removed 
with a pipet and kept in a clean bottle. This will keep for 1 or 2 weeks. 

The stain consists of: 

Alum, saturated aqueous solution, 10 c.c. and 
Gentian violet, saturated alcoholic solution, 2 c.c. 
This should not be more than 2 or 3 days old when used. 

The film prepared as above described is covered with as much of the 
mordant as the cover-glass will hold and heated for about one minute over a 



THE URINE: BACTERIOLOGY 283 

flame just hot enough so that the fluid will steam gently. It is then well 
washed for about two minutes in running water and carefully dried over a 
flame. The specimen is then covered with the stain, heated, allowed to 
steam for about a minute, washed well in water, dried and mounted. 

The smears of the sediment will give some clew as to the presence of 
organisms and as to the nature of some. In the case of certain organisms 
it is the only way we have to study them. This is especially true of the 
tubercle bacillus and the streptococci. 

Bacillus tuberculosis (for staining methods, see page 25) may be 
found in the urine in cases of tuberculosis of the kidney or of any portion 
of the genito-urinary tract, providing that the kidney is still secreting urine 
and that the tuberculous focus has ulcerated into this tract. They appear, 
however, rather late in the disease and therefore are of little value in diag- 
nosis. Bacillus tuberculosis is not infrequently found in the urine of tuber- 
culous patients with apparently normal kidneys. They have been found 
in the urine of patients with miliary tuberculosis, 165 pulmonary tuberculosis 
and in fact with tuberculosis of any organ of the body. Some teach that 
their presence in the urine indicates tuberculosis of the urinary tract only 
when pus also is present in the urine, or when there are other signs of 
local tuberculosis. 

Cases of sterile pyuria are often due to tuberculosis. There is no stain- 
ing method which absolutely differentiates tubercle bacilli and the smegma 
bacilli in urine examinations. Cultural methods and animal inoculations 
are of value if positive. The urines to be examined should be obtained by 
catheterization using most careful aseptic precautions. 

It is possible that some of the nonpathogenic acid-fast bacilli ingested 
with the food may appear in the urine but this must be rare. 

To avoid smegma bacilli Young and Churchman (see page 284) advise 
thorough cleansing of the penis, rinsing it with large quantities of water, 
and careful irrigation of the anterior urethra. The urine, they say, should 
be passed into 3 glasses and only the third used in examination for tubercle 
bacilli. This technic, they believe, will fully exclude all smegma bacilli 
from the urine and any acid- and alcohol-fast bacilli present can be con- 
sidered tubercle bacilli. In women the ureters must be catheterized. 

Petroffs Method of Examination of the Urine. — The urine to be examined 
is acidified with 30% acetic acid and then 2% of its volume of a 5% solution 
of tannic acid added. The specimen is then put in the ice-chest for 2 4 hours. 
The precipitate can then be centrifugalized, redissolved with dilute acetic 
acid, centrifugalized and the sediment placed on slides and stained; or, the 
first precipitate may be treated with normal sodium hydroxide solution 
and cultivated. 

The smegma bacilli are a group of organisms which grow in abundance 
on the external genitalia and wherever the secretions of the skin are allowed 

166 Churchman, Am. Jour. Med. Sci., July, 1905. 



284 CLINICAL DIAGNOSIS 

to accumulate. Their morphology and staining characteristics vary con- 
siderably. Some of the strains resemble so closely the tubercle bacilli both 
in morphology und in acid- and alcohol-fast staining reactions that they 
cannot be differentiated by this method. Twenty-one different stains 
have been published to differentiate these organisms from Bacillus tuber- 
culosis, but in vain. It is fortunately not necessary as a rule to try to differ- 
entiate between them, since it is much easier to avoid them entirely by 
using care in obtaining the specimens, in which case any acid-alcohol-fast 
bacilli found can safely be called Bacillus tuberculosis. 

Recent work 166 would tend to prove that smegma bacilli are difficult 
to cultivate directly from the patient. In fact, some hold that real smegma 
bacilli cannot be cultivated; others, that smegma contains two varieties 
of acid-fast organisms only one oi which can be grown. Brereton and Smith 
found in the cases of 126 insane or uncleanly patients red staining bacilli 
in 85 (67.5%) if the specimens were decolorized by 25% sulphuric acid 
and in only 19 (22%) if methylene blue was used as a counterstain after 
decolorization. They were present in 13% if the specimens were decolorized 
by acid alcohol. In a second series of 20 men of ordinary cleanly habits, 
smegma bacilli were present in 13 (65%), if the specimens were decolorized 
with sulphuric acid only (25%), and in only 2 (10%) if the smears were 
counterstained. 

Smears from the anterior urethra (fossa navicularis) of 24 patients 
showed (Young and Churchman) smegma bacilli in 11 (46%), while of 
6 patients, they were found in the urine of 5. The urine in the bladder at 
necropsy and smears from the bladder wall were negative in 50 cases. The 
posterior urethra was negative for smegma bacilli in the 6 cases examined. 

Streptococci also are much more safely searched for in smears from the 
sediment than by cultural methods. 

In conclusion, no matter what the organism is which is present, one 
should always control his cultures by a preliminary bacterioscopic 
examination. 

THE BACTE.RIOLOGY OF THE URINE, CULTURAL METHOD 

The last portion of the urine voided is well centrifugalized, or first 
diluted with sterile water to dilute the urine and to wash the sediments 
(certainly no alcohol may be used), and cultures are made from the sediment. 

The culture medium used will depend in great measure on the organisms 
suspected. When, however, the nature of the organism is not known, 
blood agar is the best medium to use, since all aerobic organisms which 
can be cultivated at all will grow on this. A few loopfuls of the urinary 
sediment are rubbed over the surface of this medium and the tubes then 
inoculated at 37 C. The different colonies can then be distinguished and 
transplantations made to suitable media. 

166 Quoted from Brown, Jour, of A. M. A., 1915, lxiv, p. 886. 



THE URINE: BACTERIOLOGY, CULTURAL METHOD 285 

The media in common use are the following : 

Nutrient Agar. — About 1500 c.c. of distilled water are heated over a 
furnace in a large metal pan while 15 gms. of agar are shredded and slowly 
added, together ith 2.5 gms. of Liebig's meat extract. The heating is 
continued, stirring at intervals until all the agar is dissolved. All floating 
scum is then skimmed off with a spoon. The pan is now removed from the 
fire, cooled slightly, and 10 gms. of peptone (Witte's) and 5 gms. of sodium 
chloride added little by little, stirring vigorously all the while to facilitate 
solution. The pan is then replaced on the fire and the contents boiled and 
stirred until all the peptone is dissolved. The fluid is then made just 
alkaline (to litmus) by the addition of a 5% solution of sodium hydrate. 
The pan is then removed from the fire and cooled to 6o° F. To it is then 
added the whites of 2 eggs which in the meanwhile have been mixed with 
150 c.c. of water. The pan is then replaced on the furnace and slowly 
heated until coagulation is complete. The fluid is not stirred while this 
coagulation is in progress. When coagulation is complete the pan and 
contents are weighed and enough water added to bring the weight of the 
contents up to just 1000 gms. (Twenty grams are allowed for the weight 
of each white of egg which will be filtered* off.) 

Meanwhile a large funnel with rubber tube and pinch-cock- on the 
nozzle has been set up on a retort stand. • In this is placed a well-moistened 
creased filter paper in a wire holder. The contents of the pan are now 
poured into the funnel through a strainer which will remove the coagulum. 
The medium is filtered at once into tubes or into a flask and sterilized for 
7 minutes in an autoclave. 

If the medium filters too slowly it is poured back into the pan, reheated, 
and then filtered through a fresh filter paper. 

This is the medium on which the most of the common organisms are 
grown. It also is the basis of other special media. 

Glycerin Agar. — This medium is similar to the above except that it 
contains from 6 to 8% of glycerin which is added after it is filtered. This 
fluid is tubed and sterilized in the autoclave for 7 minutes. It is superior 
to plain agar since many organisms which grow delicately on that grow 
well on this. This is true of streptococci, the meningococcus, the pneumo- 
coccus and the tubercle bacillus. 

Blood Agar (Rosenau's Method). — This is one of the most valuable of 
media. About 100 sterile slant tubes of plain agar are first prepared. To a 
flask containing 50 c.c. of plain agar warmed to from 40 to 6o° C, are added 
(under aseptic precautions) 15 c.c. of human blood. This is well mixed 
by shaking and from 1 to 2 c.c. are poured into each of the slant agar tubes 
which are then placed in such a position that this agar-blood mixture may 
harden as a uniform layer over the plain agar slant. In this way a little 
human blood will make a great many tubes. This technic must be aseptic 
to prevent contamination, since these tubes should not be sterilized again. 



286 CLINICAL DIAGNOSIS 

The tubes are then left in the thermostat for a few days to be sure that 
they are sterile. 

On this medium will grow practically all aerobic organisms which can 
be cultivated at all. It is especially good for the gonococcus and Bacillus 
influenzae. 

Nutrient Gelatin. — Nutrient gelatin is made in practically the same way 
as is nutrient agar except that instead of agar one uses from 106 to 150 gms. 
of " gold leaf " gelatin. This medium should, however, be boiled as little 
as possible and should be sterilized in the autoclave for not over 5 minutes. 
(Of course tubes of this medium should not be placed in a thermostat kept 
at body temperature, but kept at room temperature or in a special thermo- 
stat the temperature of which does not rise above 22 C. 

Litmus Milk. — About 100 c.c. of fresh milk are allowed to stand in the 
refrigerator for 5 hours and as much of the cream as possible removed. 
Then enough litmus tincture is added to give the milk a deep sky-blue 
color. The medium is then tubed and sterilized in the autoclave for 
7 minutes. 

Bouillon. — The formula is : 

Liebig's meat extract 2.5 gms; 
Peptone (Witte's) 10 gms; 
Sodium chloride 5 gms; 
Water, distilled, 1000 c.c. 
The steps for making this medium are practically the same as those for 
plain agar. To the boiling water is added the meat extract and the boiling 
then continued for 5 minutes. The pan is then cooled down a little, the 
peptone and salt added slowly and then dissolved by boiling and the fluid 
made alkaline as described above. This medium when finished is first 
poured into a flask and sterilized in an autoclave for 5 minutes, then cooled, 
filtered twice through the same paper, then tubed and sterilized again in 
the autoclave for 5 minutes. 

Loffler's Blood-serum Mixture (see page 37). 

Among the more important organisms which may be encountered in 
the urine are the following: 

Bacillus Coli Communis. — The colon group includes 20 or 30 very 
similar varieties which usually are grouped under this one name. It is a 
short organism, the majority from 1 to 2/* long and o.$ijl thick. Some, 
however, are so short as to resemble cocci and others are over 5/1 long. 
They often are seen in pairs. The most of the varieties are sluggishly 
motile, although this may not be evident in cultures over 24 hours old, 
while other strains are very motile. Its flagella are numerous and laterally 
placed. It is easily stained by all the usual aniline dyes and is decolorized 
by the Gram method. It does not produce spores. The organisms with 
unstained portions resembling spores are involution forms. It grows 
rapidly on all ordinary media at room as well as at body temperature and 



THE URINE: BACTERIOLOGY, CULTURAL METHOD 287 

in a characteristic way. The growth on the surface of agar is abundant, 
thick, moist and spreads rapidly. The deep colonies are very circumscribed 
and opaque, with a well developed nucleus. This bacillus does not liquefy 
gelatin and does not spread on the surface of this medium as it does on 
agar. The surface colonies on gelatin have a nucleus and well-defined 
granular or striated, refractive halo. It turns litmus milk rapidly acid 
(within 1 8 hours), coagulates it (in from 4 to 30 days) and does not later 
digest the clot. The growth on potato is abundant and visible. It ferments 
almost every known carbohydrate, but especially glucose, lactose and 
saccharose and with abundant gas production. It produces indol. The 
colon bacillus is almost ubiquitous. It is the prevailing organism of the 
lower part of the small intestine and the colon and is almost universal in 
soil, water, food, etc. It is a mildly pathogenic and pyogenic organism. 
It is the organism found most frequently in infections of the urinary tract. 

Bacillus Typhosus. — The typhoid bacillus is a long slender organism 
measuring usually from 2 to 4/1 in length and about 0.5^ in thickness. It 
is actively motilar. Its flagella are more numerous (from 5 to 50) and some- 
what longer than those of Bacillus coli communis. It is easily stained by 
all the aniline dyes and decolorizes by Gram's method. It does not produce 
spores. It grows on all ordinary media. The growth on agar is thin and 
translucent with slightly spreading dentate or leaf -like edges. The deep 
colonies have sharply defined edges and a distinct nucleus. Bacillus typho- 
sus does not liquefy gelatin. Its growth on potato is often, but by no means 
always, invisible. (The colon bacillus grows on potato as a distinct brown- 
ish scum and the typhoid bacillus may also. This seems to depend on the 
potato used.) A most important feature is the reaction of this bacillus to 
litmus milk. A very slight acidity is first produced, distinct, but never 
enough to coagulate the milk not even in weeks. This slight acidity may 
be permanent although some strains later furnish enough alkali to change 
the reaction back to neutral, or even to alkaline. Certain carbohydrates, 
including dextrose, levulose, maltose and mannites are fermented to the 
point of acidity, but none with gas production. Saccharose and lactose 
are not at all affected. It does not form indol. 

For the certain recognition of this bacillus its agglutination in the very 
dilute serum (1 : 50 to 1 : 1000) of a patient or animal immune to Bacillus 
typhosus is necessary (see page 565). 

This organism is present in the urine of % of all cases (see page 291) of 
typhoid fever during the fever and in some cases for years afterward. 

The Paratyphosus Group of Organisms. — The more carefully the 
cases of " typhoid fever " are studied bacteriologically the more numerous 
those are found to be which are due not to Bacillus typhosus but to very 
similar bacilli which resemble also, in some essential respects, Bacillus coli 
communis. Those which stand nearest this latter organism are called 
" paracolon "bacilli, those nearer Eberth's bacillus, " paratyphoid." There 



288 CLINICAL DIAGNOSIS 

is no one Bacillus paratyphosus, but a group. In the clinical laboratories 
at least eleven such organisms have been isolated, although for all practical 
purposes of diagnosis we recognize but 2: Bacillus paratyphosus A, and 
Bacillus paratyphosus B. The court of last appeal in the recognition of 
these bacilli is the serum agglutination test, using the serum of an animal 
immunized to the organism in question. This is fairly satisfactory although 
they show a rather marked group reaction. Clinically we note whether or 
not the patient's blood will agglutinate one of them. 

The characteristic culture features of the paratyphoid bacilli are their 
reaction to milk and to sugars. All resemble Bacillus typhosus in that they 
produce first a slight acidity in milk, but they all (and this may take weeks) 
finally change the reaction back to alkaline. The attempt to divide the 
group into the "A" group of organisms which keep the milk acid and the 
"F" group which change it back to alkaline seems unjustified. 167 Again, 
these all ferment glucose with gas production. Some ferment lactose, also 
some saccharose, but none all of these three sugars. 

Bacillus Lactis Aerogenes. — Bacillus lactis aerogenes is a short, 
thick (about 2/i thick) non-motile, encapsulated bacillus, seen usually in 
pairs and sometimes in chains. It often shows rather marked polar stain- 
ing. It is decolorized by Gram's method. It is not a spore-producing 
organism. It produces on all media a luxuriant, viscid, slimy growth which 
resembles in many ways that of Bacillus coli communis. It does not 
liquefy gelatin. It ferments practically all the carbohydrates rapidly and 
with abundant gas production. This organism has been the subject of 
much dispute. Some do not distinguish it from Bacillus coli communis, 
some group it with the capsulated group. Some strains of this organism 
cannot be distinguished from Bacillus capsulatus of Friedlander (Bacillus 
pneumonias) . 

This bacillus is a normal inhabitant of the upper part of the small 
intestine where it is the predominating organism. Some believe it the 
cause of certain cases of cystitis. 

Bacillus Alkaligenes. — This bacillus is a normal inhabitant of the 
intestinal tract and has been often mistaken for Bacillus typhosus. It is 
a long, slender bacillus from 2 to 3/z long and 0.5^ thick, which grows 
singly, in pairs, or in chains and which is actively motile. 

It grows on all culture media. It does not liquefy gelatin. Its most 
characteristic cultural reaction is the production of an intense alkaline 
reaction in litmus milk without previous acid production. It ferments 
carbohydrates without producing an acid reaction and without gas produc- 
tion and is the only one of the common intestinal flora which grows only in 
the open end of the fermentation tubes and not at all in the closed end. 

The Proteus Group. — The members of this group, among which are 
Proteus vulgaris, Proteus mirabilis, Proteus zenkeri, Proteus zoffii, and 

167 Ford, Medical News, June 17, 1905. 



THE URINE: BACTERIOLOGY, CULTURAL METHOD 289 

other strains with but minor differences, are the most important agents of 
putrefaction. While secondary invaders as a rule, they are in certain cases 
of cystitis the pathogenic organism. In fact this is almost the only organism 
which when introduced into a normal bladder will set up a cystitis. They 
are the most important organisms in the production of a " sapremia," or 
intoxication from the products of decomposition, as in cases of retained 
placenta. They are short, slender, actively (even violently) motile bacilli 
with terminal flagella. 

This description of the organism applies only to those obtained from 
fresh cultures. If cultures a little older are examined one will find in the 
smears cocci, bacilli of all sizes, spirilla, etc., suggesting a badly contam- 
inated culture. But if fresh transplantations be made from this culture 
and examined early the organisms will be found to be bacilli of uniform 
morphology. This marked polymorphism is due to the tendency of this 
organism to produce involution forms. It is decolorized (?) by Gram's 
method. It grows well at room temperature. The colonies on agar spread 
rapidly in a characteristic manner since the edges send out peculiar hair- 
like projections which make the colonies look like tufts of moss. Proteus 
vulgaris and mirabilis liquefy gelatin (the former rapidly, the latter slowly) , 
the blood -serum rapidly; Proteus zenkeri and zoffii do not. Milk is coagu- 
lated and clot then digested, the reaction remaining all the while alkaline. 
The strains of the proteus groups differ somewhat in their ability to ferment 
sugars. The formerly much-talked-of Bacterium termo is possibly one of 
this or of a nearly related group. 

Bacillus Pyocyaneus. — Bacillus pyocyaneus is a small organism 
about 2 fi long and 0.5/x thick, actively motile, which is easily stained by all 
ordinary bacterial stains and which shows often a marked bipolar staining. 
It decolorizes by Gram's method. It grows rapidly on all ordinary media 
and will crowd out any other organism which happens to grow along with 
it. Its important cultural characteristics are that it will not split up 
carbohydrates, that it liquefies gelatin and blood-serum rapidly, that it 
coagulates litmus milk rapidly, decolorizing the litmus and then digesting 
the milk clot with the reaction alkaline and, lastly, that it produces 
2 pigments in its growth — a non-specific fluorescent pigment and a specific 
bluish-green pigment called " pyocyanin." 

This is a very common organism often met with in the intestine and on 
the skin, especially in the folds of the axillae and groin. It is the pyogenic 
bacillus which produces blue pus; it sometimes is the organism of septi- 
cemia of children ; but its most important role as a pathogenic organism is 
as the cause of cystitis and ascending genito-urinary infections. 

Bacillus Aerogenes Capsulatus. — This is one of the most widely 
distributed of pathogenic organisms. It is a constant inhabitant of the 
intestine of man and of animals and is commonly present in the soil, water, 
milk, etc. 
19 



290 CLINICAL DIAGNOSIS 

This is a large bacillus, from 1.5 to 6/x long and ifx thick. It is non- 
motile, is encapsulated and produces spores in the animal body and when 
grown on blood-serum. It is easily stained by all the ordinary stains and 
does not decolorize by Gram's method. 

It is a pure anerobe, growing only in the complete absence of oxygen. 
It grows best (under anerobic conditions) at 37 C. in the depth of solid 
media as grayish-white or brownish colonies with fine feathery or hair-like 
projections from their edges. It ferments sugars easily. Litmus milk is 
coagulated, decolorized and the clot later digested. 

Since it is met with very often in mixed infections the best way to 
cultivate it is to heat the material containing the mixture of organisms at 
86° C. for a few minutes to kill off all but the spores. Plate cultures are 
then made and cultivated anerobically. The most of the organisms in 
such a mixture are not spore producers and so will be killed off leaving 
but a few forms alive of which this will be one. 

A still better way to isolate this organism is to inoculate a rabbit intra- 
venously with the material containing the mixture of organisms. After 
5 minutes the animal is killed and placed in the thermostat for from 6 to 
8 hours or left in a room temperature for 18 to 24 hours. The animal will 
during this time become much distended with gas. Bacillus aerogenes 
capsulatus can now be obtained from the blood in almost pure culture. 

This organism is one of the most important of the pathogenic bacteria. 
Its infections are extremely grave. Those of the genito-urinary tract are 
not infrequently the result of dirty technic in obstetrics. 

Bacillus Tetani. — The tetanus bacillus is a slender organism about 
4 or sn long and 0.5/z thick. It is a motile bacillus with a great many very 
long, slender flagella. It is a spore-producing bacillus and since the spores 
have a diameter 3 or 4 times that of the bacillus and are usually situated 
at the end of the bacillus the drum-stick shape of the sporulated organism 
is characteristic. Those with a spore at each end resemble a dumb-bell. 
These spores are very resistant to heat and are not killed by a temperature 
of 8o° C. for 1 hour. 

This bacillus is easily stained by all the ordinary bacterial stains, the 
body of the bacillus taking an unusually uniform stain. It is not decolorized 
by Gram's method. 

The tetanus bacillus is a perfect anerobe and can be isolated only with 
extreme difficulty. Fortunately it is not necessary to grow it since its 
appearance in smears is quite characteristic. 

It is one of the most ubiquitous and important of pathogenic organisms. 
Its normal habitat would seem to be the intestine of cattle and so may be 
found wherever the soil is contaminated with manure. 

The bacteriological study of the urine is often important in the diag- 
nosis of septicemia, acute nephritis, pyelitis, ureteritis, cystitis, prostatitis 
and urethritis. In case of septicemia one would try first to isolate the 



THE URINE: BACTERIOLOGY, CULTURAL METHOD 291 

organism from the blood. Since the importance of bacilluria in the spread 
of diseases has been recognized, the presence of organisms in the urine has 
aroused a new interest. It is now known that Bacillus typhosus can be 
found in the urine of about one-third of the cases of typhoid fever during 
the attack. Sometimes these bacilli are few in number, sometimes so 
numerous, even 500,000,000 in each cubic centimeter, that they actually 
cloud the fresh urine. This bacilluria may clear up with the fever but it 
also may persist for years without giving rise to any local symptoms 
which might warn these "chronic bacillus carriers" that they are spreading 
this disease far and wide. 

In other forms of septicemia also the invading organism is often found 
in the urine. The presence in the urine of Bacillus tuberculosis in cases 
of chronic tuberculosis, especially of the acute miliary form, has already 
been mentioned (see page 283). In some cases of streptococcus infection 
the fresh urine on the day of death is turbid because of the streptococci 
it contains. 

Infectious Nephritis. — So many cases of acute nephritis date to an 
acute infectious disease, such as pneumonia, tonsillitis, influenza, typhoid 
fever, etc. (to say nothing of scarlet fever, measles, etc. — diseases the spe- 
cific organisms of which are not yet known) or to an acute infection, as in 
a recent case of streptococcus infection of the arm complicated by acute 
nephritis, otitis media leading to meningitis, etc., that it is not unreasonable 
to believe that the nephritis is due to the same organism as that which 
causes the primary infection and that cultures from the urine therefore 
may help discover this germ. Especially may this be true of the interesting 
cases of unilateral nephritis. 

Acute pyelitis is due to an infection which usually reaches the pelvis 
of the kidney from the blood stream but which theoretically may be an 
ascending infection secondary to a cystitis, or possibly one which has 
traveled along the lymphatics. Fortunately now it is an easy matter to 
catheterize the ureters of patients and so get urine for cultures directly 
from the pelvis of each kidney. In a pyuria of renal origin if the 
cultures are sterile the cause is usually tuberculosis of the kidney. We 
will speak of the bacteriological findings in pyelitis in connection with those 
in cystitis. 

It may be mentioned at this point that a pyelitis without localizing 
symptoms is quite common and that a pyelitis may continue for consider- 
able time before a cystitis begins. 

Cystitis. — Cystitis is an inflammation of the urinary bladder due to 
some pathogenic organism. This bladder when normal is very resistant 
to infection and organisms introduced into it soon disappear unless some 
predisposing factor has lowered the resistance of its wall. The 2 exceptions 
to this general rule are Bacillus tuberculosis and proteus, which if intro- 
duced into a normal bladder can set up a cystitis. Among the conditions 



292 CLINICAL DIAGNOSIS 

favoring infection may be mentioned: calculus ; posterior gonorrheal 
urethritis ; frequent catheterizations ; the retention of urine from any cause 
(very important) such as childbirth, enlarged prostate, urethral stricture, 
spinal cord disease and prolonged narcosis; and, finally, infection of the 
rectum with secondary involvement of the bladder. 

Tuberculous Cystitis. — Among the forms of cystitis the cases due to 
tuberculosis form so well-defined a group that they deserve special mention. 
A primary tuberculous cystitis is a rare condition. This infection usually 
descends from the kidney or ascends from the genital tract. The descend- 
ing cases are particularly important since all the symptoms often are vesical 
and the renal disease, even though of extreme grade, may be unsuspected 
until cystoscopic examination suggests a higher origin and ureteral cathe- 
terization determines the source and nature of the pus. 

Tuberculous cystitis is a disease especially of young adults. It begins 
insidiously and may last for years. There is usually in well-developed 
cases increased frequency of micturition, but not always. The urine is 
acid and contains pus, often considerable, a condition which may have 
escaped the patient's attention. Cultures made from it are sterile. There 
are 2 general rules of value. All persistant, acid pyurias in the young are 
presumably tuberculous until the contrary is proven (Kelly); and all 
sterile (in ordinary cultures) pyurias are either tuberculous or gonorrheal 
in nature (see page 294). It is easier to demonstrate the tubercle bacillus 
(see page 283) in tuberculous cases than the gonococcus in those cases. 

There is an interesting group of very early cases of tuberculous cystitis 
with practically no pus at all in the urine, with slightly increased frequency 
of micturition and slight, transitory hematuria, which may persist for a 
few days and then not reappear for weeks. The urine is clear .and rather 
highly colored, but the last few cubic centimeters of the voiding usually 
contain a few drops of blood. 

In advanced cases of tuberculous cystitis the urine remains acid, the 
pyuria is marked, tubercle bacilli are easy to demonstrate and there is 
persistent or recurring hematuria. Following a secondary infection, and 
sooner or later this is quite certain to develop, the pus usually increases in 
amount and the urine often becomes alkaline. 

It is important to remember that cases of tuberculous prostatitis and 
ureteritis may so resemble tuberculous cystitis that only a cystoscopic 
examination will localize the lesion. 

Cystitis due to Organisms other than Bacillus Tuberculosis. — Among the 
organisms found in cystitis are Bacillus coli communis, Staphylococcus 
aureus, Staphlycoccus albus, Streptococcus pyogenes, Bacillus proteus 
vulgaris, Bacillus pyocyaneus, Bacillus typhosus, Bacillus lactis aerogenes 
and others. In many cases it is difficult to determine just how important 
in the etiology of the cystitis is the organism isolated in cultures. Many 
are certainly secondary invaders and harmless saprophytes. 



THE URINE: BACTERIOLOGY, CULTURAL METHOD 293 

Bacillus coli communis is the most important of the above list. It 
causes a very chronic cystitis. 

It must be very difficult indeed for the gonococcus to gain a primary 
foothold in the bladder, for in practically every one of the very numerous 
cases of gonorrheal posterior urethritis this organism must frequently 
enter the bladder. The region of the trigone would seem the most suscepti- 
ble spot in the bladder to this infection, which once developed may be 
quite stubborn. Other cases of definite gonorrheal cystititis develop as 
part of an acute urethritis and clear up rapidly. The gonorrheal and, 
tuberculous infections are the only 2 which seem exceptions to the general 
rule that pyuria is an almost invariable symptom of cystitis. It is very 
difficult to demonstrate the gonococcus in these cases, and secondary 
pyogenic infections, a not uncommon sequel of gonorrheal cystitis, usually 
mask the picture. 

Proteus cystitis is a common and very distressing form. Bacillus 
proteus seems to be almost the only organism which when introduced into 
a normal bladder will set up a cystitis (Melchoir). The urine in these cases 
is very alkaline and the abundant pus is transformed into a ropy, sticky, 
mucoid mass. 

Streptococcus cystititis is often a very severe form, but sometimes is 
mild. Bacillus lactis aerogenes is believed to be a much commoner cause 
of cystitis than statistics would lead one to expect. 

The catheterized urine of cases of cystitis practically always contains 
pus. Certain cases of tuberculosis and of gonorrhea are exceptions to this 
rule. As mentioned above the kidney should always be excluded as the 
source of much of this pus, especially if there is much fever or if there is 
more albumin than the pus alone would explain. As a rule the most of the 
pus appears in the first and the least in the second of the 3 -glass test, but 
this holds true only if the patient had been resting before voiding. If the 
reaction of the urine is acid the pus-cells often are well preserved, sometimes 
ameboid, and settle as a granular layer on the bottom of the glass; when 
very alkaline, the pus is transformed into a sticky, ropy, mucoid mass. 

The reaction of the urine in the tuberculous cases is usually acid until 
secondary infection by proteus or a streptococcus occurs. In chronic 
cystititis due to the colon bacillus, Staphylococcus albus and other organ- 
isms with slight virulence, the urine may be either acid or alkaline. In 
the proteus and streptococcus cases the urine is alkaline, the phosphates 
are precipitated, the pus-cells unrecognizable and the odor of the urine foul. 

Red blood-cells are numerous in the urine of cases of acute cystitis, 
their number varying with the acuteness of the attack. They are uniformly 
distributed throughout the urine. The hemorrhages from the bladder wall 
are slight. The larger hemorrhages come from the kidney, from vesical 
tumors, or especially from the prostatic urethra. In cases of posterior 
urethritis the blood may continuously ooze back into the bladder and when 



294 CLINICAL DIAGNOSIS 

these patients void the urine is at first bloody and at the end of the voiding 
pure blood. 

For mention of epithelial cells in the urine in cystitis, see page 262. 

In "membranous," "exfoliative," " croupous," "diphtheritic," or 
" desquamative " cystitis the patient passes flakes, masses, or moulds of a 
tough fibrinous membrane containing much degenerated epithelium. These 
masses are supposed to be the results of necrosis of the inner layers of the 
bladder wall. In gangrenous cystitis fragments of the epithelial and mus- 
cular coats of the bladder are expelled in the urine. In hemorrhagic cystitis 
there may be much bloody infiltration of the bladder wall. Clinically these 
severe forms of cystitis are very rare. They occur often enough after the 
traumatic or operative opening of the bladder and as terminal events. 

Bacteriuria. — When a bacteriuria is present the urine when voided may 
contain so many organisms that it is actually cloudy. The symptoms of 
these cases often suggest a severe cystitis and yet cystoscopic examination 
may show the bladder normal, although later a mild cystitis is almost 
certain to develop. 

A transitory bacteriuria often follows massage of the prostate gland. 
It begins within a few hours after this procedure and may last 1 or 2 days. 
There are no symptoms. These organisms are supposed to come originally 
from the rectum. 

The cases of persistent bacteriuria fall into two groups. The first is 
of renal origin, already mentioned on page 291, and due usually to Bacillus 
typhosus, Bacillus coli, etc. The second group included those cases which 
are secondary to a posterior urethritis and prostatitis. While the foci of 
infection are in these organs the organisms multiply in the bladder and 
are so numerous that they may cloud the urine. Very little pus is present 
in the urine of these patients, 

In gonococcus, typhoid, and colon bacteriuria the urine remains acid; 
in streptococcus and staphylococcus bacteriuria its reaction may be acid, 
neutral, or alkaline ; in the Staphylococcus albus bacteriuria, however, the 
urine is usually alkaline. 

INFECTIONS OF THE URETHRA AND EXTERNAL GENITAL ORGANS 

In this connection it will be necessary to describe 3 very important 
organisms not mentioned in the preceding pages. 

The Gonococcus. — The gonococcus is so important an organism that 
the medical student should be skilled in its recognition. Formerly supposed 
to be the cause of unimportant, transitory, local infections, it is now known 
to be one of the most destructive of organisms. 

The gonococcus (see Fig. 60) is a coccus about i/jl in diameter, which 
occurs, as a rule, in pairs. The proximal edges of these diplococci are 
flattened, giving the organism the well-known biscuit shape. In smears 
of gonorrheal pus the organisms are found chiefly inside pus and epithelial 



THE URINE: INFECTIONS, URETHRA AND GENITALIA 295 



cells, or in masses on their surface; but some are free. It stains well in all 
the ordinary bacterial stains, but especially in methylene blue, and dis- 
colorizes by Gram's method. (This organism in smears of pus, however, 
decolorizes so slowly that it may be necessary to leave the specimen in 
alcohol for 10 minutes.) The gonococcus grows only on media containing 
some human proteid, as human blood agar and blood-serum, but agar 
mixed with ascitic fluid or hydrocele fluid will be satisfactory, or a medium 
made up of urine, blood-serum and agar. Its growth on proper media is 
a thin, moist homogeneous layer which dies in a few days. Cultivated 
under the best of conditions and transplanted frequently, the gonococcus 
will survive but for a few generations. It is very susceptible to changes in 
temperature. It dies in a few hours in pus at room temperature and is 
quickly killed by a temperature of 40 or 41 C. It is killed rapidly by 
drying. For its recognition it is neces- 
sary to assure oneself that it occurs as 
biscuit-shaped diplococci, in clusters 
or clumps, some of them at least in- 
tracellular, that at least 1 cell found 
contains many such organisms, that 
it decolorizes by Gram's methods, 
and that it grows only on the above- 
mentioned media. 

The gonococcus is the important 
causative agent of urethritis and 
periurethritis, prostatitis and infec- 
tion of the connecting ducts and 
glands of the genito-urinary system, 
the seminal vesicle, prostate, epidid- 
ymis, bladder, Bartholin's glands, vagina, uterus, tubes, etc. ; it causes an 
eczematous skin eruption about the genitals; it is an important cause of 
proctitis, peritonitis, meningitis, endocarditis and especially of conjunctiv- 
itis, arthritis and septicemia. A definite discharge is not a necessary 
indication of possible gonorrheal infection, for it may produce very little 
pus, as in the vaginitis of infants and in chronic arthritis. 

Much has been written of organisms often found in the normal urethra 
which morphologically " are exactly similar to the gonococcus, and which 
also decolorize by Gram's method." These, however, will grow easily on 
ordinary culture media. One would not be likely to mistake these if he re- 
members that the gonococcus is found in groups or clumps of organisms in a 
smear, while one would find but 1 or 2 of the pseudogonococci on a slide. 

Acute Anterior Urethritis. — The discharge in a case of acute urethritis 
during the first few hours is scanty, resembles dilute milk or starch solution 
and consists chiefly of serum, epithelial cells, a very few leucocytes, often a 
few red blood-cells and a few gonococci, the most of which are extra- 




Fig. 60. 



Spread of pus containing gonococci. 
(Wilson.) 



296 CLINICAL DIAGNOSIS 

cellular. After a few hours, however, the discharge becomes more abund- 
ant, yellow, creamy in consistency and is then an almost pure bloody pus 
in which gonococci are easily found. During the first very few days of 
the infection the gonococcus will be the only organism found, but later 
the ordinary rich urethral flora returns. 

In an untreated case, which may clear up spontaneously in from 4 to 6 
weeks, the discharge again becomes starchy, more and more scanty and con- 
sists then of abundant mucus with fewer pus-cells, but with more and more 
epithelial cells. The gonococci meanwhile become fewer and more difficult 
to find. Finally the discharge is almost pure mucus which contains no pus 
or gonococci. In case the disease had not extended beyond the anterior 
urethra the patient is now well, but only too often the infection extends to 
the posterior urethra and adjacent structures, in which case secondary infec- 
tions by pyogenic organisms are common and these modify the exudates. 

In posterior urethritis the discharge is often profuse, but since it is re- 
strained by the compressor urethra muscle it all must flow back into the 
bladder and be voided with the urine. The discharge from the anterior 
urethra may at this time be very scanty. If this patient passes his urine 
in two portions the discharge then present in both posterior and anterior 
urethra will be washed out with the first portion of urine while the second 
glass will contain the pus which has been flowing back into the bladder; 
yet this second glass should not contain as much pus as does the first glass. 
If the amount of exudate from the posterior portion be small the second 
specimen of urine may be clear. If the anterior urethra is first well irrigated 
with boric acid solution and then the patient voids into 2 glasses, the pres- 
ence of pus in the first will indicate a posterior urethritis. The best time 
to try this test is with the first voiding in the morning. 

The sequelae of posterior urethritis are prostatitis, vesiculitis, epididy- 
mitis and cystitis. In very acute posterior urethritis the frequent and 
excessively painful micturition is a very distressing symptom. The whole 
urine may be colored by the blood which is constantly flowing back into 
the bladder and at the end of micturition a few drops or more of pure 
blood often flow from the urethra (" terminal hematuria"). 

If a chronic urethritis follows an acute, the discharge may be continu- 
ous and fairly profuse or very scanty. In the latter case there may be 
only enough exudate to glue together the lips of the urethral orifice but as a 
rule there is a little exudate which consists of shreds of mucus enclosing a 
few pus, but more epithelial, cells. This discharge should be carefully 
distinguished from the glairy discharge which follows an acute urethritis. 
Smears should be carefully studied for pus-cells and gonococci. This 
exudate is washed out of the urethra as Tripperfaden (see page 273). It 
is very difficult to demonstrate the gonococcus in such an exudate. 

These " clap threads " when long, translucent and branching are made 
up mainly of mucus which is washed out of folds in the urethral mucosa ; 



THE URINE: INFECTIONS, URETHRA AND GENITALIA 297 

others are short, thick, tack-shaped and sink quickly to the bottom ot the 
glass. These contain considerable pus. They are supposed to come from 
the urethral crypts. Some of the shreds from the posterior urethra are 
short, slender, delicate, and comma-shaped. These are from the prostatic 
excretory ducts (Furbinger's hooks). These shreds should be carefully 
examined for gonococci. 

In other cases of chronic urethritis the discharge is an abundant, thick 
pus in which pyogenic organisms also may be present, but not always. 
Still other cases have an intermittent discharge of muco-pus or almost 
pure mucus. 

If the patient immediately after irrigating the anterior urethra voids 
into 3 glasses and shreds are found in any of the glasses there surely is 
present a chronic posterior urethritis. In these cases the whole volume of 
urine may be slightly cloudy because of the exudate which is constantly 
flowing back into the bladder. 

In women an acute urethritis is of briefer duration and less apt to 
become chronic than in men. (Many deny this.) Next to the urethra, 
the cervix in women is the most common focus of gonorrheal infection. 
The cervical discharge is at first slimy and blood-stained and later a milky 
pus. In this location, especially, is the infection apt to become latent and 
chronic, and its only sign a viscid catarrhal mucous discharge. 

The discharge in cases of vulvitis and vaginitis is, because of secondary 
infections, often a profuse and very fetid pus. 

Non-specific Urethritis. — Not all cases of urethritis are due to the gono- 
coccus. Some are due to various other organisms, the colon bacillus, the 
diphtheria bacillus, streptococci, staphylococci, etc. 

Smears of the exudate of the cases not gonorrheal in nature will show 
the presence of these other organisms in great numbers. Nevertheless, 
it is well to remember the frequency with which secondary infections com- 
plicate a true gonorrhea and the difficulty one often has to find the gono- 
coccus when it is present. 

In these non-specific cases the exudate is purulent, but not profuse and 
the cases are mild and respond readily to treatment. 

Bacteriorrhfca. — Some patients have as urethral discharge a thin opaque 
fluid, which microscopically consists almost entirely of mucus and sapro- 
phytic bacteria of all varieties, but no pus-cells. These patients have no 
other symptoms. The discharge clears up quickly under treatment. This 
condition is called bacteriorrhea. 

Prostatitis. — Prostatitis may be due to the extension of an infection 
from the urethra or from the rectum, or to an hematogenous infection, 

The diagnosis of acute prostatitis is more a matter of physical than 
urinary examination. In chronic prostatitis 168 the diagnosis is made by 

168 p or complete description see Young, Johns Hopkins Hosp. Reports, 1906, vol. 
xiii, p. 302. 



298 



CLINICAL DIAGNOSIS 




Pig. 6i. — Prostatic fluid. (X 400.) a, 
lecithin globules ; b, pus-cells; c, epi- 
thelial cells; d, corpora amylacea; e, free 
granules from epithelial cells ; /, sper- 
matozoa. 



physical examination but also by the examination of the prostatic fluid 
which one obtains by massaging the prostate gland after the urethra has 

been well irrigated. 

The normal prostatic fluid, which is a 
thin, bluish, skim-milk-like fluid, is de- 
scribed on page 3 08 (Fig. 61). The fluid in 
a case of chronic prostatitis may contain 
none, some, or all, of the normal consti- 
tuents of this fluid and in varying amounts. 
No diagnostic or prognostic value can as 
yet be ascribed to the presence or relative 
amounts of these normal constituents. The 
abnormal element of greatest importance is 
pus. In some cases at times, and especially 
on the first examination, no pus-cells will be 
found and yet later, many. The amount of 
pus present bears a fairly direct relation to 
the extent of prostatic involvement. Red cells are sometimes abundant. 
Spermatozoa, active or immobile, are found in varying numbers. In 
chronic prostatitis the fluid is always alkaline to litmus. 

When there is so 
much prostatic fluid that 
a discharge follows uri- 
nation or defecation the 
condition is called "pro- 
statorrhea" (also called 
"spermatorrhea"). This 
usually indicates a rather 
mild prostatitis. 

The fluid which can 
be expressed from the 
seminal vesicles is a 
thick and gelatinous se- 
cretion which resembles 
boiled tapioca or sago. 
This fluid sinks in the 
urine. Its chief constit- 
uents are " mucin glo- 
bules," large unformed 
masses resembling large non-nucleated epithelial cells (Fig. 62, b), sper- 
matozoa (Fig. 63 represents a large mucin globule full of spermatozoa), 
pigmented epithelial cells, and small finely and coarsely granular non- 
nucleated epithelial cells. Some cells might be mistaken for casts (see 
Fig. 64). 



-&©?& 




b 



J- o c A" vv o O 



Sec 



FtG. 62. — Prostatic fluid: a, epithelial cells; b, clear epithelial cells, 

from seminal vesicles (?) ; c, corpus amylaceum; d, "granular cells " 

with droplets resembling myelin; e, "granular cells" with fat 

droplets. 



THE URINE: INFECTIONS, URETHRA AND GENITALIA 299 

For purposes of instruction the students may well use the 7 -glass test in the differ- 
ential diagnosis of chronic inflammatory lesions of the urethra, prostate, seminal vesicles 
and bladder, although this test is not often used in practice. The patient compresses 
the urethra far back at the root of the penis (at the suspensory ligament) while the 
anterior urethra is irrigated by means of a long irrigating tube. The fluid is caught in 




Fig. 63. — Mass of mucus filled with spermatozoa from urine catheterized at death. X 400. 

2 glasses. The first, I \ will contain the shreds, if any are present, the second, I 2 , should 
be perfectly clear. The patient's fingers are then removed and the tube carried back 
as far as the deeper part of the bulbous urethra. The washing is again caught in 2 glasses. 
The first, 1 3 , will contain the shreds from the bulbous urethra (if any are there), the 
second, 1 4 , should be clear. The urine is then voided into 3 glasses, I 5 , 1 6 , 




Fig. 64. — Cells which resemble casts found in fluid messaged from 
a prostate, the seat of chronic prostatis. (Kindness of Dr. George 
Walker, of Baltimore, who will later publish this and similar cases.) 



1 7 , which will contain bladder urine, and mixed uniformly in each the exudate of 
posterior urethritis which has flowed back into the bladder between voidings. In addi- 
tion to this: 

I 5 will contain the exudate in the posterior urethra which will usually be washed 
clean by the flow of urine. 

1 6 may contain the last traces from the posterior urethra, and 

1 7 will contain also urine from the most dependent portions of the bladder, also the 
contents of the prostatic and ejaculatory ducts which often do not discharge the exudate 
collected in them until the muscular contractions made at the end of micturition force 
out the plugs of thickened exudate which occlude their mouths (Ftirbinger's hooks). 



300 CLINICAL DIAGNOSIS 

MICRO-ORGANISMS OF THE EXTERNAL GENITALIA 

Bacillus Ulceris Cancrosi (Ducrey's Bacillus). — This organism is 
now recognized as the cause of soft chancre. It is found in smears of the 
purulent discharge from these sores, but always mixed with a host of other 
organisms. (Sections of the tissue show it in pure culture.) 

This bacillus is a small oval rod about 1.5/x long and 0.5/z thick, which 
stains readily in all bacterial stains, but decolorizes very easily. It is a 
very poor grower indeed but some claim it can be cultivated on blood agar. 

Treponema Pallidum, Spirocheta Pallidum. — This organism (see Fig. 
65) is a spirocheta the average length of which is from 6 to 15/x (although 
some are even 20^ long), and thickness 0.25^. Noguchi 169 described 3 
strains which differ in coarseness: the thicker, 0.3/x in width; the thinner, 
0.2/*, and the average, 0.25^. It is tightly twisted like a corkscrew in 5 
or 6 or more regular, closely set and rigid curves, the fineness and regularity 
of which is a characteristic feature. It is pointed at each end. It is a 
flagellated organism, but shows only a twisting, rotating, or bending 
motion. They are usually found singly, although sometimes several 
are tangled. 

To secure a good specimen when searching for Treponema pallidum 
one should choose a lesion which is likely to be rich in these organisms, as 
the suspected chancre, mucous patches, tonsils, condylomata and enlarged 
glands. It is also possible to obtain them from a fresh skin eruption. In 
the case of superficial lesions the serum should be obtained from the deeper 
layers of tissue, since this usually is richer in treponema and freer from 
contamination than is that from nearer the surface. Among the con- 
taminating spirochetal are : in the mouth, Spirocheta bucallis, Treponema 
microdentium and macrodentium, Treponema mucosum, and the Spi- 
rillum of Vincent; on, or near the genitals, Spirocheta refringens and 
Treponema calligyrum. 

The examination of sores and ulcers which may be chancres is espe- 
cially important because the Wassermann test at this stage of lues may 
be negative. 

The sore to be examined is first washed with sterile physiological salt 
solution, then its superficial tissue is removed by means of a sharp, sterile 
curet. The blood is wiped away with a piece of clean gauze. After the 
bleeding lessens and >the blood becomes more serous, the sore is squeezed 
by the fingers and the drop of blood which is pressed from the deeper tissues 
is caught on a clean thin slide and at once covered with a cover-glass if the 
fresh specimen is to be examined with dark-field illumination. The cover- 
glass is gently pressed down to secure a thin, even layer of serum, drops of 
cedar oil are applied to the cover-glass and another opposite this on the 
slide and the specimen then fitted onto the stage of a microscope supplied 

160 Jour. Exp. Med., 1912, vol. xv, p. 201. 



THE URINE: MICRO-ORGANISMS OF GENITALIA 301 

with attachments for dark-field work. The light must be carefully manipu- 
lated or the thin, delicate treponema may be overlooked. This method 
of examination is the surest as well as the quickest and easiest. The stain- 
ing methods are slow and less reliable. In primary and secondary lues this 
is a safer method than the serum reaction. 

If a medicated dusting powder, an ointment, or a wash has been used 
on the lesion, the search usually is fruitless. In that case the lesion should 
be washed for several days with plain soap and water and then the exami- 
nation made. 

Enlarged lymph-glands may be carefully aspirated with a fine needle 
on a tightly fitting glass syringe. 

In examining mouth lesions great care should be taken to obtain the 
serum from the deeper layers of tissue, since so many other spirochetal 
may be present. 

In the examination of skin lesions the superficial layer of epidermis is 
removed by means of a sharp curet and the serum squeezed from the cutis. 

To find spirochetas similar in appearance from quite different parts of 
the body will often clear up a doubtful diagnosis. 

If blood is to be examined, i c.c. of blood is mixed with 10 c.c. of 0.3% 
acetic acid, this fluid is then centrifugalized and the sediment examined. 

If stained specimens are desired very thin smears should be made. 
This organism is stained with great difficulty. 

The Giemsa staining mixture in common use is : 



Azur II eosin, 3 gms. ; 

Azur II, 0.8 gm.; 

Glycerin (Merck C. P.), 250 gms.; 

Methyl alcohol (Kahlbaum I), 250 gms. 

The specimen, dried in the air and then fixed for 1 hour in absolute alcohol, 
is stained for 24 hours in a fresh dilution of this stain (1 drop of the above 
mixture to 1 c.c. of distilled water). The specimen is then examined. If 
the nuclei of the leucocytes have taken a deep blackish red color the smear 
is well enough stained to justify the long search necessary to find the 
organisms, which take a delicate violet-purple color. 

Cultivation of Treponema Pallidum. 170 — A piece of sterile fresh 
rabbit's kidney or testicle is placed at the bottom of each of 6 tubes which 
measure 20 cm. in length and 2 cm. in width and then into each tube is 
poured 15 c.c. of mixture consisting of 2 parts of 2% agar, slightly alkaline 
and heated to 50 C, and 1 part of ascitic or hydrocele fluid. After this 
solidifies a layer of sterile paraffine oil 3 cm. deep is superimposed. 

The tissue from which cultures are to be made is first cleansed with 
sterile salt solution and then suitable fragments are snipped off. These, 
are immediately immersed in sterile salt solution containing 1% sodium 

no Noguchi, Jour, of Exp. Med., 191 1, xiv, p. 99 and 1912, xv, p. 90. 



302 CLINICAL DIAGNOSIS 

citrate. These fragments of tissue, which should be carefully preserved 
against drying, are then cut into small bits. One such bit is emulsified in 
a mortar with the citrate solution and this emulsion examined under the 
dark-field microscope to determine definitely that spirochetal are present. 
The other bits of tissue are used to inoculate the tubes of media. One of 
these particles is forced to the bottom of each culture tube by means of a 
blunt glass rod or heavy platinum loop, and into the same tube several 
drops of the emulsion are introduced deeply by a capillary pipet, care 
being taken to avoid tearing the medium. 

These tubes are inoculated at 37 C. continuously for 2 or 3 weeks 
before examining them. By that time they usually show a dense opaque 
growth of bacteria along the stab-canal and the diffuse opalescence of 
spirochetal growth radiating from this. The solid medium is now pierced 
by a capillary pipet to obtain some of this latter growth which is examined 
under the dark-field. A mixed culture is generally found. Fresh tubes 
are now inoculated with material obtained with a capillary pipet, which 
is introduced deeply into the agar column even to the bottom of the tube, 
without striking the central stab-canal. The contents of the pipet are 
expressed by means of compressed air, as, for instance with a syringe. 
These tubes are inoculated for 2 or 3 weeks at 37 C. and are again inspected. 
If the faint hazy zone around the stab-canal (and due to the growth of 
spirochetas) has extended far enough to allow one to penetrate this zone 
with a capillary pipet one makes another transfer as follows : 

The surface of the medium is sterilized with sublimate alcohol. The 
tube is now scratched about its middle with a diamond pencil and the glass 
cracked by a red-hot glass rod which is touched to the scratch. The upper 
half of the test-tube is then removed. The exposed surface of the agar 
is sterilized with sublimate alcohol, after which all remaining moisture is 
wiped away with sterile absorbent gauze. The agar column is now bent 
gently until it cracks transversely thus exposing a clear surface on which 
the hazy growth is readily seen. Now, without touching the central canal, 
a capillary pipet is introduced into this haze and, after confirming by 
dark-field examination the presence of pallidum, a fresh series of inocula- 
tions is made. Several reinoculations are as a rule necessary before a pure 
culture is obtained. 

Treponema pallidum is very anerobic and migrates into the solid media 
from the inoculation stab. This organism can be grown in liquid media. 171 

SpirochetaRefringens (see Fig. 65, d). — Spirocheta refringens is often 
found together with Spirocheta pallidum. It is a common parasite occur- 
ring in great numbers in many ulcerative lesions. It is larger, thicker and 
more refractile than Treponema pallidum. Its spirals are broader, more 
wavy and more irregular, its ends are blunter, it occurs in greater numbers 
in a smear and it stains more easily than does pallidum. 

171 Noguchi, Jour, of Exp. Med., 1912, xvi, p. 211. 



THE URINE: MICRO-ORGANISMS OF GENITALIA 303 

Treponema Microdentium. — Noguchi 172 was the first to isolate Tre- 
ponema microdentium in pure culture from the mouth. It is obtained from 
tooth deposit, preferably from a young child, and grown in much the same 
manner as is Treponema pallidum. It is, however, a much more rapid 
grower. This organism measures less than 0.2 $fj, in width at the middle of 
the body and gradually tapers towards both extremities, which are sharply 
pointed. Organisms from old cultures may reach S/jl in length and show, 
on an average, 14 curves. The ends are usually drawn out straight. It is 
non-pathogenic. The cultures have a characteristic putrefactive odor, not, 
however, that of pyorrhea alveolaris. 

Treponema Macrodentium. — Noguchi 173 found Treponema macro- 
dentium most frequently in the mucus about the tonsils and pharynx and 



5 






: 






O 


* ^ 


■ 


L , 




C— 




Fig. 65. — On the left Treponema pallidum (Spirocheta pallidum). A smear 

from a chancre. On the right Spirocheta refringens. A smear from a 

chancroid. 

in large numbers in the exudate in ulcerative stomatitis. Its cultivation 
is more tedious than that of microdentium. Its morphology differs con- 
siderably according to the age of the cultures. Young organisms are plump 
and short with from 2 to 8 rather irregular shallow curves. Its width varies 
from 0.7 to i.ofj,, and its length from 3 to S/jl. It is doubly refractile. Its 
cultures have no odor. 

Treponema Mucosum. — Noguchi 174 isolated Treponema mucosum 
from the pus around the roots of the teeth of a case of pyorrhea alveolaris. 
Its most striking features are its capacity to produce ip pure culture a 
mucin and a strong fetid odor. The pus was gathered from the gum 
margin in a sterile capillary pipet and suspended in a few cubic centi- 

172 Jour, of Exp. Med., 1912, xv, p. 81 

173 Ibid. 

m Jour. Exo. Med., 1912, xvi p. 194. 



304 



CLINICAL DIAGNOSIS 



meters of sterile citrate solution. Tubes of medium similar to that used 
for Treponema pallidum are innoculated with this suspension culture. 

These organisms measure from o.25too.3^in width, and from .8 to 12/1 
in length. The curves number from 6 to 8, are remarkably regular and 



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'"•> ' ' s '::'; ; f| - — ^ 






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Fig. 66. — Protophytes and other low forms of life often found in tap water. 

X 400. 

often quite deep. Both extremities are sharply pointed. It cannot be 
distinguished from Treponema pallidum by its morphology alone, but can 
by cultivation. This organism explains in part at least the characteristic 
odor of the breath in pyorrhea alveolaris. 

Treponema Calligyrum. 175 — Treponema calligyrum is an organism 
found by Noguchi on the surface of genital and anal lesions of luetic andnon- 



175 Noguchi, Jour, of Exp. Med., Jan. 1. 1913. 



THE URINE: YEASTS AND MOULDS 



305 



luetic patients. It is non-pathogenic. Morphologically it stands between 
Treponema pallidum and Spirocheta refringens. It is easily cultivated. 
On the skin about the genitalia is an abundant flora of organisms: 
streptococci, staphylococci (especially Staphylococcus albus), Bacillus coli 
communis, Bacillus pyocyaneus, Bacillus lactis aerogenes, Bacillus aero- 
genes capsulatus, and various strains of smegma bacilli. 




FlG. 67. — Biastomycetes in the urine. 
YEASTS AND MOULDS IN URINE 

The protophytes added from tap water should be recogn'zed (Fig. 66). 

Yeasts. — Ordinary yeast cells from the air or surface of the body often 
reach the urine and sometimes multiply so abundantly that they are con- 
spicuous in the sediment, especially in cases of diabetes. They may gain 
entrance to the bladder and there ferment the sugar before the urine is 
voided thus giving rise to " pneumaturia." If they are to be cultivated 
the urine should be kept acid with acetic acid. 

In cases of systemic blastomycosis this organism may be found in the 
urine in large numbers (see Fig. 67). 
20 



306 CLINICAL DIAGNOSIS 

Moulds may occasionally be found in fresh urine but the most are later 
contaminations. In one case of pyelitis, previously treated by repeated 
irrigations through ureteral catheters, the urine on one occasion was very 
bloody, and contained masses of the mycelium of some mould which would 
not grow on the various media tried. One thorough irrigation cleared up 
this infection. The chances are that at a previous catheterization some 
spores were introduced into the pelvis of the kidney. 

Sarcinas, smaller than those found in the gastric contents, may also be 
found in the urine. 

ANIMAL PARASITES 

Among the animal parasites which may be demonstrated in the urine 
are the hooklets, daughter cysts (even several hundred in a case) and frag- 
ments of membrane of echinococcus cysts. There will be no urinary 
symptoms of hydatid disease of the kidney, unless perhaps a catarrhal 
pyelitis, unless the cyst ruptures into the urinary tract and then the urine 
will appear watery, soapy, or bloody. Embryos of filaria are found in the 
urine in cases of tropical hematochyluria (see page 209) . 



FlG. 68. — Eggs of Eustrongylus gigas. X 400. 

Flagellates belonging to the cercomonas or trichomonas groups often 
are found in the urine. There is a dispute whether they have ever been 
found in a fresh catheterized specimen. Many may be later contamina- 
tions. Balantidium coli however would seem to infect the urinary tract 
and be found in the freshly voided urine, from 5 to 15 parasites in each 
field under the microscope, as in Hinkelmann's 176 case. 

Eustrongylus Gigas. — A few cases of Eustrongylus gigas infection 
have been reported, but in most of them there certainly was a mistake in 
diagnosis. In 1 case of chyluria, however, the eggs were found 177 (see 
Fig. 68). 

Schistosoma Hematobium (Bilharzia). — The trematode worm, Schis- 
tosoma hematobium (see Fig. 69), so common in Africa, especially in 
Egypt and the Transvaal, has been found twice in Porto Rico (Martinez) 
and six times in this country. 178 

The male measures from 12 to 14 mm. in length, is flat but so folded 
that it appears cylindrical and forms a gynecophoric canal which receives 

176 New York Med. Jour., Jan. 30, 1915. 

177 Stuertz, Deutsches Arch. f. klin. Med., 1903, vol. lxxviii, p. 586. 

178 See O'Neil, Boston Med. and Surg. Jour., October 27, 1904, vol. cli, p. 453, also 
Dr. Daywalt's letter in Dr. Arnold's paper, Southern Practitioner, 1906, vol. xxviii, p. 13. 



THE URINE: ANIMAL PARASITES 



307 




: v.v 



**%: 



the female. The female is 20 mm. long and is filiform. The adults live 
in the portal vein, also its branches and in other veins of the abdomen and 
pelvis, especially those of the bladder, the pelvis of the kidney and the 
rectum. The eggs (Figs. 102 and 103) are large, from 120 to 190^ long and 
from 50 to 73/x wide, are fusiform, have no operculum and have a spine 
which may be terminal or lateral (Schistosoma mansoni ?) . 

The urinary symptoms of this 
infection are catarrh of the blad- 
der and hemorrhages ("Egyptian 
hematuria"). At first only blood 
flecks are passed at the end of 
micturition but later the hemor- 
rhages may be profuse . The symp- 
toms are caused by the eggs 
which the female deposits in the 
mucous membrane. These may 
be passed with the urine, but any, 
especially those with a lateral 
spine, remain in the bladder and 
form the nucleus of calculi. Each 
egg contains a miracidium which 
is completely ciliated. The shell 
splits when the urine is diluted 
with water. What the inter- 
mediary host is, and how human 
infection occurs, are not known. 

Nematode worms other than 
filaria are sometimes found in the 
urine, especially Anguillula aceti, 
or the "vinegar eel." Stiles re- 
ports one case of infection of the 
bladder with this worm. Other 
cases may be due to contamina- 
tion from the bottle in which 
the urine is collected. 179 These 
worms resemble closely Strongyloides intestinalis, except that they 
(A. aceti) are slightly longer (males 1.2 mm. long and 0.033 ram- 
wide; females 1.9 mm. long and 0.06 mm. wide; embryos 0.25 to 0.3 
mm. by 0.015 mm.). 

The student should always be able to recognize the various plant con- 
taminations which occur in tap and stagnant water and in vessels rinsed 
out with this water. That is, he should be able to recognize that they are 
of no significance (see Fig. 66). 

179 Billings and Miller, American Medicine, May 31, 1902. 




Fig. 69. 



Schistosoma hematobium, adult worms. 
(Copied from Braun.) 



308 CLINICAL DIAGNOSIS 



PROSTATIC FLUID 



The prostatic fluid is best obtained by " milking " this gland by the 
finger in the rectum. The urethra is first well washed, then the fluid thus 
expressed and collected. The amount under normal conditions varies 
greatly, from none to even 5 c.c. at 1 milking. It has a grayish- white, 
yellow, or greenish color, a milky turbidity due to lecithin globules and a 
characteristic odor. It is slightly viscid, tenacious, of low specific gravity 
and contains but little solid matter (from 1 to 2%). The reaction of the 
prostatic fluid has attracted considerable attention because of the possi- 
bility that this may be an important factor in the production of sterility. 
Thus far no results of importance have been gained. It reacts faintly 
alkaline to most reagents and acid to others, but this varies much. 

One examines this fluid for motile spermatozoa immediately on obtain- 
ing it, then adds a drop of acetic acid to bring out the cells more clearly 
and examines it for pus-cells (Fig. 61, b). 

Microscopically, the most striking objects in prostatic fluids are the 
great numbers of lecithin globules (Fig. 61, a) which give it its milky 
appearance. These vary in size from those very minute to others even 
half the size of a red blood-cell. They are not very refractive and so can 
be distinguished readily from fat. An increase in their numbers in the 
fluid of patients with chronic prostatis indicates improvement. Corpora 
amylacea (Fig. 61, d; 62, c) are sometimes found in the urine, especially 
of elderly men. They have no significance. They are laminated and have 
a finely granular center. Of their composition nothing is known except 
that they stain blue with iodine. Epithelial cells of various kinds are 
present. Some are large and polygonal, others are cylindrical; they are 
found single or in groups, and vary much in size (see Figs. 61, c; 62, a). 
Interesting cell-like masses which vary much in size, the so-called granular 
cells (Fig. 62, e), which may resemble colostrum corpuscles, are merely 
masses of fat-like granules. These break down and liberate the refractive 
globules seen free in the fluid (Fig. 61, e) . Some of the granules resemble 
myelin (Fig. 62, d). Columnar epithelial cells and cylindrical cells which 
resemble casts (see Fig. 64) are sometimes present. One finds also large 
clear cells of varying size, with or without a nucleus (see Fig. 62, b), which 
are supposed also to arise in the seminal vesicles. In normal prostatic 
fluid one finds no pus-cells and no red blood-corpuscles. Spermatozoa 
(Fig. 61, /) are usually present in prostatic fluid. 180 

Prostatic fluid should be examined for spermatozoa while it is as fresh 
as possible since it is important if possible to see them move. To study 
their finer structure the fluid, if much proteid is present, should be diluted 
with even 20 volumes of water and smears made which are dried in the 
air, heated to 120 and cooled slowly. The specimen is then covered with 

180 For a description of these, see University of Pennsylvania Med. Bull., No. 3, 1902. 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 309 

2% iron-alum solution for from 2 to 4 hours, washed in water and then 
with 1% hematoxylin for 12 hours. It is then decolorized carefully with 
1% iron-alum, counterstained for from 1 to 3 minutes with a saturated 
aqueous solution of eosin, dried and mounted. Many of the spermatozoa 
are abnormal in shape. Some have 2 heads and some even 3 tails. These 
monsters are never seen to move. One seldom tries to determine more 
than their presence and the motility. If they are motile, one concludes 
that they are functionally normal. If they are not found or are not moving 
no conclusions at all are justified. 

In fluids from cases of acute or chronic prostatitis many leucocytes 
are present and the lecithin globules are diminished in number. 

Spermin crystals may be demonstrated by adding to the prostatic 
fluid 1 drop of 1% ammonium phosphate solution and allowing the speci- 
men to dry for 2 hours under the cover-glass. These crystals resemble 
somewhat the Charcot-Leyden crystals. They are colorless, transparent 
needles or whetstones, many of which are imperfectly crystalized. 

Prostatic casts or testicular casts (see page 273). 

Gonorrheal threads are described on page 296. 

The short coma -like flocculi which are sometimes seen, form in the excre- 
tory ducts of the urethral glands and follicles and mean an intense involve- 
ment of these structures. Those found in the second glass are from the 
prostatic glands and are signs of chronic prostatitis. They consist of super- 
imposed layers of cylindrical epithelium. 

Prostatic plugs, which are large cylindrical masses of mucus, are some- 
times seen. These form in mild inflammations of the prostatic ducts. 
Some of these mucous masses are full of spermatozoa (Fig. 63). 

FUNCTIONAL RENAL DIAGNOSIS 

Methods of estimating with some degree of accuracy the functional 
ability of various organs and especially of the kidneys is one of the best 
recent contributions to medicine. The discrepancies between the kidneys 
as found at autopsy and that which one would expect from the urinary 
examinations alone are proverbial. Small contracted kidneys, less than 
% or % the normal size, may excrete a urine normal in amount and in specific 
gravity, which contains but a trace of albumin and very few casts ; in other 
cases with marked albuminuria and cylindruria no clear evidence of 
nephritis may be found ; while some patients have died in so-called uremia 
who had little clinical evidence of renal insufficiency, whose urine contained 
but a trace of albumin and whose kidneys at autopsy showed slight changes. 
Evidently the time-honored clinical, chemical and microscopical methods 
are no test of the important lesions. They may indicate the severity of 
the acute lesion then in progress, but they certainly do not test the amount 
of renal substance left by disease, nor the ability of this to do its work 
(see page 323). Surely there must be some methods of predicting an immi- 



310 CLINICAL DIAGNOSIS 

nent death from renal insufficiency, some means of foreseeing an on-com- 
ing uremia. These means may be found in functional renal diagnosis and 
in blood chemistry. 

The problem of functional renal diagnosis is not so much to determine 
the anatomical condition of the kidney as its ability to do a stated amount 
of work in a given time. A well-compensated severe lesion is manifestly 
of less immediate danger than a poorly or non-compensated acute lesion. 

Before describing the tests for renal functional ability it is well to state 
that the toxins so evident from their results in nephritis are themselves 
as yet unknown unless it be that they are the same bacteria which produce 
infections elsewhere in the body. We believe this to be true and yet the 
kidney function is not reduced in every general infection nor yet in many, 
as a study of the general septicemias shows, while in many cases of severe 
or fatal nephritis the evidence of general infections may be unsatisfactory. 
It is easy to assume that the kidney must be a relatively difficult organ to 
infect since its function is to remove all poisons from the body and this 
would include the toxin of infections. It is also easy to assume that a 
case of nephritis is chronic because its cause is chronic. • At least we can 
say that there is little evidence of the vicious circle formerly so empha- 
sized which was supposed to keep a nephritis in progress, for once clean 
out the infections elsewhere in the body and in some cases the nephritis 
at once begins to subside. The function tests impose upon the kidneys a 
definite task and measure their success in fulfilling it. From their success 
in eliminating a given amount of some substance in a given time we by 
analogy surmise how well they perform their other functions. At the 
onset it should be confessed that the functions of the kidney are not as 
well understood by the physiologist as the clinician seems to assume when 
he uses the methylene-blue test to test the " epithelial filter," the salicylic 
acid test to test the "glomerular filter " and phlorizin to test the " glandular 
activity " of the renal epithelium. 

No one test is above criticism; one is never enough and several must 
be used to get a fair renal picture. 

We wish to warn workers that these functional tests make an unusual 
demand on the kindeys and one to which they may not be able to respond, 
so that slight reactions do follow and disastrous results may follow even 
the simplest. 

Physiochernical Tests — Cryoscopy, Freezing Point of the Urine. — 
Cryoscopy, or the determination of the depression of the freezing point 
of water by a salt in solution, is a well recognized and very valuable method 
of physical chemistry to determine the molecular weight of a substance 
and the degree of disassociation of its molecules. Clinical chemists at- 
tempted (by determining the freezing point of urine and of the plasma) to 
estimate how many products of katabolism the kidneys have excreted in 
the urine and how many are still in the plasma. But even when the prob- 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 311 

lems are of the simplest nature, i.e., when dilute solutions of a single and 
pure salt are used, this method requires experience, skill and due regard 
to a good many factors which can modify the results. It is hard to see, 
therefore, how this method could be applied with much success to complex 
fluids like the urine or blood, in which are dissolved a great variety of 
bodies of widely different nature, some unknown; and yet most definite 
conclusions concerning the functional ability of the kidney were drawn 
from changes in the freezing point of these fluids which a physical chemist 
would consider slight. 

Electrical Conductivity. — This excellent method of physical chem- 
istry also was appropriated by the clinician in his desire to estimate the 
functional ability of the kidneys. By electrical conductivity is meant the 
reciprocal of the resistance which a certain amount of a solution between 
2 platinum electrodes of given size and given distance apart offers to the 
passage of a current of known strength. This is really a measure of the 
number of electrolytes in solution, that is, of the disassociated ions. It 
is not affected by such bodies as albumin, sugar, urea, which are not dis- 
associated, and hence is practically a measure of a few salts in the blood 
and urine, especially the chlorides. It is difficult enough to get accurate 
results using a simple dilute solution of one salt and it certainly is working 
in darkness to apply this very delicate method to the blood and urine, 
which contain an unknown mixture of various bodies. It therefore is little 
wonder that this method gave nothing definite. 

Methods Using Normal Products of Metabolism. — The time of excre- 
tion of urea is an old criterion of functional renal ability. In acute nephritis 
it may take 3 to 6 days to excrete the urea resulting from 1 day's meals; 
in chronic nephritis and renal tuberculosis, 2 days. One therefore gives 
the patient a meal of measured ingredients and then follows the curve of 
the elimination of urea. During this delay the urea accumulates in part 
in the blood and may be determined there quantitatively (see page 542). 
When it is increased tenfold {i.e. , to 0.3%) there is danger of uremia (Herter) . 

The Salt and Water Test. — The patient is kept for a few days on 
a diet constant in water (about 2000 c.c. per day) and in chlorides (about 
5 gms. per day, all the chlorides of foods included) . After 2 days on which 
the chloride output has been fairly constant the patient is given at 1 time 
from 5 to 10 gms. of salt and the curve of its elimination determined. 

Normally, an increased salt intake is eliminated in 1 of 2 ways, depend- 
ing on the amount of water given at the same time. If given without 
increasing the water intake it is excreted within 24 hours and without any 
diuresis. If extra water was given at the same time its excretion is accom- 
plished partly by increasing the concentration of the urine and partly by 
diuresis. The excretion of salt is by the tubules (Schlayer). 

In cases with moderately severe vascular injury of the kidney the 
administration of the salt may be followed by a marked diuresis and all 



312 CLINICAL DIAGNOSIS 

the salt be excreted within 24 hours, but without any increase in the con- 
centration of the urine (i.e., its specific gravity remains rather low and 
fairly constant). This condition (vascular hyposthenuria of Schlayer) 
seems due not so much to a lesion of the tubules as to hypersensitiveness 
of the vessels. If, however, the vascular lesion is severe the kidney responds 
to the administration of the salt by an oliguria. 

In cases with severe tubular destruction the urine is little affected by 
the dose of extra sodium chloride, since the tubules are unable to excrete 
it (tubular hyposthenuria). 

The Dilution Test. — The excretion of water is so important a func- 
tion of the kidney that it is natural to assume that variations in this would 
be a very simple and satisfactory test of renal permeability. This test is 
applied in a simple form as follows : The patient drinks an unusual amount 
of water with his evening meal and practically none until the following 
morning. He voids on retiring, which should be at least 4 hours after the 
meal, and again in the morning. A normal person will eliminate the most 
of the excess of fluid in the evening specimen which will therefore have a 
lower specific gravity than the morning specimen. If the specific gravity 
of the morning specimen is lower than that of the evening there is evidence 
of vascular renal lesion; but if the 2 are almost equal this " fixation of 
specific gravity " indicates a severe grade of renal insufficiency. 

In a more elaborate way this test may be performed as follows: the 
specific gravity of the patient's urine is determined and then he is asked 
to drink from 1 to 2 liters of water. The time of the appearance and the 
duration of the resulting polyuria and of the lowering of the specific gravity 
are noted. To determine the combined efficiency of the kidneys each half 
hour's voiding is examined separately but to determine the relative effici- 
ency of the 2 kidneys the ureteral catheters are introduced and left in place 
for at least 3 hours and the urine from each is collected in half -hourly portions. 

The dilution of the urine may begin during the second half hour, may 
reach its maximum in 2 or 3 hours and last 5 or 6 hours. In parenchymatous 
nephritis the ability to eliminate an excess of water seems more reduced 
than in cases of contracted kidney. If the kidneys are not equally diseased 
the excretion of the diseased one will be more uniform than that of the 
more normal organ; that is, the more normal kidney will show the dilution 
first and more markedly that the other. 

The test thus refined has little value. Even in the normal person we 
find no uniformity in the time of the appearance of, or in the duration of, 
a polyuria following the ingestion of large amounts of water. Again, 
there may already be a polyuria of the diseased side which will mask any 
increase of the output of the more normal side. And, lastly, 3 hours is a 
long time to allow ureteral catheters to remain in position. 

Test Meals. — Several test-meals have been proposed to test renal 
function. That is, the patient consumes weighed meals selected with a 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 313 

view to taxing somewhat the renal function and the urine is studied to 
ascertain the ability of the kidneys to meet the problem. 

Hedinger and Schlayer had proposed as an improvement over v. Mona- 
kow's " added urea and salt test," which requires 10 or 12 or even more 
days, a 2 -hour renal test which could be completed, so far as the patient is 
concerned, in 24 hours. The results are very much the same. Mosenthal 181 
proposed the modification of their diet described below. All food is to be 
salt-free food from the diet kitchen. Salt for each meal will be furnished 
in weighed amounts. All food or fluid not taken must be weighed or meas- 
ured after meals. No other food or fluid of any kind is allowed. 

Breakfast, 8 a.m.: 

Boiled oatmeal 100 gms. 

Sugar 1-2 teaspoonfuls. 

Milk 30 c.c. 

Two slices of bread (30 gms. each). 

Butter 20 gms. 

Coffee 160 c.c. 

Sugar 1 teaspoonful 200 c.c. 

Milk 40 c.c. 

Milk 200 c.c. 

Water 200 c.c. 

Dinner, 12 noon: 

Meat soup 180 c.c. 

Beaf steak 100 gms. 

Potato (baked, mashed or boiled) 130 gms. 

Green vegetables as desired. 

Two slices of bread (30 gms. each). 

Butter 20 gms. 

Tea 180 c.c. 

Sugar 1 teaspoonful [2 00 c.c. 

Milk 20 c.c. 

Water 250 c.c. 

Pudding (tapioca or rice) no gms. 

Supper, 5 p.m.: 

Two eggs cooked in any style. 
Two slices bread (30 gms. each). 
Butter 20 gms. 
Tea 180 c.c. 

Sugar 1 teaspoonful [2 00 c.c. 
Milk 20 c.c. 

Fruit (stewed or fresh) 1 portion. 
Waterloo c.c. 
181 Arch, of Int. Med., 1915, xvi, p. 733. 



314 CLINICAL DIAGNOSIS 

No food or fluid is to be given during the preceding night or until 
8 o'clock the next morning (after voiding) , when the regular diet is resumed. 

The patient empties his bladder at 8 a.m. and then each 2 hours until 
8 p.m. The urine from 8 p.m. to 8 a.m. is collected in 1 specimen. 

The above diet contains approximately 13.4 gms. of nitrogen, 8.5 gms. 
of salt, 1760 c.c. of fluid and a considerable quantity of purin material 
in the meat, soup, tea and coffee which act as diuretics. 

'Hare's Modification of Hedinger and Schlayer's " Two-hour Test. 182 

O'Hare, the next year, proposed the following menu for the test day: 

Seven a.m.: coffee, milk, sugar, toast, and butter; 10 a.m.: milk, 
toast and butter; 12.30 p.m. : bouillon, broiled steak, butter, mashed potato, 
butter, toast and butter, coffee, milk and sugar; 4 p.m.: tea, milk, sugar, 
crackers; 7 p.m.: soft egg, blanc mange (1 egg, sugar, cornstarch, milk) 
and cream. Amounts sufficient to give approximately 2500 calories, 
1550 c.c. of fluid, 76 gms. of protein, 127 gms. of fat, 245 gms. of carbo- 
hydrates and 5.8 gms. of sodium chloride. 

This is a mixed diet containing food diuretics of various types (purins, 
salts, water, etc.). Each of the 5 unequal portions contain known but 
varying amounts of fluid, nitrogen and salt — the noon meal the most 
of all. 

On the 2 days preceding this day the patient should receive about 2000 
calories, 75 gms. of protein and 4 gms. of sodium chloride. The urine is 
collected each 2 hours from 7 a.m. to 9 p.m. and then 1 " night specimen " 
from 9 p.m. and 7 a.m. Each specimen is analyzed for volume, specific 
gravity, total nitrogen concentration, total chloride and chloride concen- 
tration and the results are charted. 

The purpose of these tests is to find out to what extent and in what 
manner the diseased kidney under stimulation by the different diuretics 
taken in the food reacts by putting out the varying amounts of water, 
nitrogen and chloride ingested. Normal cases respond by putting out water 
and salt promptly and in good amounts. The diseased kidney, however, 
may show that its power to excrete is too low to accommodate the large 
amounts of these elements ingested, especially at the noon meal. These 
" fixed " cases show a curve of excretion that approaches more or less a 
straight line. Instead of getting the normal " picket fence " curves we 
find some or all of the curves of excretion more or less flattened out. 

In general the salt excretion is impaired before there is much disturb- 
ance of water and nitrogen excretion; in most patients salt and water 
excretion behave very similarly ; the nitrogen excretion is greatly impaired 
only in the severe cases. Salt, water and nitrogen excretion, however, 
show some disturbance in mild cases in which the phenolsulphonephthalein 
test is normal and there is no increased blood nitrogen. These dietary 
tests cannot be used in very severe cases of chronic nephritis. 

182 Arch, of Int. Med., June, 1916, xvii, p. 711. 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 315 

The specific gravity (quoted from Mosenthal) of the different voidings 
of a normal individual varies 10 points or more from the highest to the 
lowest. The night urine has a high specific gravity (1.018 or more), a 
high percentage of nitrogen (above 1%) and is small in amount (400 c.c. 
or less) . The quantities of water, salt and nitrogen excreted approximate 
the intake. When kidney function is lowered the first signs are usually 
demonstrated in the night urine; its quantity increases while its specific 
gravity and nitrogen concentration are lowered. One or all of these 
departures from the normal may occur. In severe cases of chronic 
nephritis the functional renal inadequacy is indicated by a markedly 
fixed and low specific gravity, a diminished output of both salt and 
nitrogen, a tendency to total polyuria and a night urine showing an 
increased volume, a low specific gravity and a low nitrogen concentra- 
tion. Such functional pictures, however, are not confined to nephritis, 
but are found regularly in many other conditions; pyelitis, cystitis, 
hypertrophied prostate, marked anemia, pyelonephritis, polycystic kidney 
and diabetes insipidus. 

The causes of diminished renal function must be sought for in the 
urinary passages, in the blood or in the kidney itself. Prognosis and therapy 
will depend largely on the cause of the fundamental impairment and not 
on its degree. A divergence between the degree of functional renal involve- 
ment and the intensity of the signs and symptoms of nephritis is frequently 
found and accentuate the lack of parallelism that there may be between 
functional and anatomical lesion. 

In chronic diffuse (parenchymatous) nephritis the condition of renal 
function is characterized by its variability. In these cases the results of 
the test -meal have proved to be extremely valuable in giving an idea of 
the status of salt, nitrogen and water excretion, as well as a picture of renal 
efficiency as a whole. The findings in myocardial insufficiency vary accord- 
ing to the condition of the heart. They differ much in the periods when 
there is myocardial decompensation and the accumulation of edema, when 
the edema is subsiding and later when cardiac compensation is again fully 
established. In such cases it requires some time before the kidney resumes 
its normal activity. This intervening period before the function is normal 
is indicated by a tendency to a low, fixed specific gravity and a nocturnal 
polyuria. During the period of full myocardial decompensation the results 
of kidney activity are very characteristic ; the specific gravity is markedly 
fixed at the level of about 1.020, the salt output is diminished, that of 
nitrogen is high in marked contrast to the salt and there is oliguria. When 
chronic nephritis and cardiac decomposition coexist, as they so often do 
in hypertensive nephritis, the urine may exhibit the characteristics 01 
either lesion. The determining factor is probably the chronic nephritis 
which may or may not be so far advanced as to present an unchanging 
barrier to the influence of renal congestion. 



316 CLINICAL DIAGNOSIS 

Ambard's Coefficient. 183 — Ambard was one of the first to show that 
the excretion of urea, sodium chloride, etc., is carried on by the kidneys 
according to definite laws capable of numerical expression. In the case of 
urea Ambard's formula, known as Ambard's coefficient, is as follows: 

Ur 



Ur = grams of urea per i liter of blood ; 

D = grams of urea excreted per 24 hours; 

C = grams of urea per 1 liter of urine ; 
Wt = weight of patient in kilos ; 

70 (kilos) is accepted as a standard weight and 25 (grams per liter) as a 
standard concentration of urea. 

The value obtained for C in normal subjects lies between 0.060 and 
0.080, the mean about 0.080.. 

Changes in C indicate changes in the blood-urea, which changes vary 
as the square root of changes in the rate of excretion. In order that patho- 
logical variations in rate of excretion may be expressed according to a scale 
of 100 (so that e.g., if the index Were 50 it would indicate that the condition 
was 50% of normal), and granting that Ambard's coefficient of 0.080 is 
the mean normal index, the formula according to McLean, 184 may be 
written as follows : 



T - _ grams of urea per 24 hours Vgrams of urea 1 L. of urine x 8.96 
Wt in kilos X (grams of urea per 1 L. of blood) 
Ambard's coefhcienc for chloride excretion. This formula as given by 
McLean is : 



Plasma NaCl = 5.62 + 



/Grams NaCl per 24 hrs. \/grams per 1 L. of urine 
* 4.23 XWt in kilos 

In this formula 5.62 is the normal threshold limit experimentally determined 
of NaCl in the plasma (e.g. no NaCl will be eliminated in the urine unless 
that m the blood exceeds 5.62 gms. per 1 L. of plasma. 

Normal Excretion of Urea and Sodium Chloride (McLean) — 
Collection of Specimens. — Short periods, e.g., of 72 minutes, during either 
the forenoon or afternoon, are preferable for observation of the excretion 
of urea and sodium chloride. By collecting the urine of a given period and 
withdrawing the blood at the middle of this period the blood sample may 
be assumed to represent the average for that period. If no food or water 
is taken during the period, and this begins not too soon after a heavy meal, 
the rate of excretion during the period will remain practically constant. 
One-half hour before the test begins the patient drinks 150 to 200 c.c. of 

183 McLean, Jour. Exp. Med., 1915, xxii, p. 212. 

184 Ibid. 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 317 

water, and takes no more fluid or food until the end of the observation. 
At the beginning of the period the bladder is emptied; 36 minutes later 
about 10 c.c. of blood are taken from an arm vein into a dry tube containing 
about 100 mgms. of powdered potassium oxalate to prevent clotting. At 
the end of 72 minutes after the bladder was first emptied the specimen of 
urine is collected, carefully measured and used for analysis. A 72 minute 
period is chosen since it is Ko of 24 hours, and the calculation for 24 hours 
is made somewhat easier. It should be remembered that the estimated 
rate of excretion for 24 hours need bear no relation to the amount actually 
excreted in 24 hours. The rate is actually determined for the shorter period 
and calculated for 24 hours as a standard period on which to base all results. 

Very accurate analyses are necessary if one desires to determine a 
quantitative relationship such as is here described. The urea content of 
the whole blood (the urea content of whole blood is slightly lower than 
that of plasma; whole blood is always used) and of the urine are deter- 
mined by the urease method. These urea determinations are always 
corrected by determinations of the preformed ammonia, the amount of 
which is subtracted. 

After the portion of whole blood for the urea determination has been 
removed the remainder is at once (within 1 hour) centrifugalized at high 
speed to throw down all corpuscles and the plasma is pipeted off (serum 
of clotted blood is never used). The total chlorides of both plasma and 
urine are then determined and calculated as sodium chloride. 

The values obtained from the urea and chloride determinations are 
substituted in the proper formulas as described. 

Water is administered before the period in order to prevent apparent 
retention due to dehydration of the organism. If a fair amount of urine 
is thus obtained the results in normal individuals will not simulate those 
of subjects actually retaining urea. Apparently sodium chloride is less 
dependent on water intake than urea. Diet, especially as regards chloride 
and nitrogen intake, is unimportant from the standpoint of the observa- 
tions as the formulas are independent of the intake. It is therefore unneces- 
sary to put an individual on a standard weighed diet in order to obtain 
comparable observations. 

The substitution in the formulas of values found by analysis and the 
calculation of the formulas is in itself a considerable task if the ordinary 
arithmetical processes are used. Logarithms are of advantage, but they 
are laborious. To simplify the process of calculation a slide-rule has been 
adapted to the formulas. By the use of this device it is not even necessary 
to remember the formulas ; the whole calculation becomes a matter of only 
a few seconds and is purely mechanical. 185 

The normal concentration of urea in the blood varies in the same or 
different normal individuals from about 0.200 to 0.500 gms. per liter. The 

185 See McLean, ibid. 



318 CLINICAL DIAGNOSIS 

rate of excretion is determined by this concentration and by the rate of 
water output. It is somewhat less for concentrations below 0.300. The 
usual range of concentration of chlorides in normal human plasma is from 
5.62 to 6.25 gms. of sodium chloride per liter or higher, according to the 
amount ingested. The rate of excretion depends on the excess over a 
threshold of about 5.62 gms. per liter. A concentration below 5.62 gms. per 
liter has not been observed in a normal individual. 

Renal Permeability to Foreign Substances. — The Methylene blue 
test of Achard and Castaigne was supposed to test the ' ' epithelial filtra- 
tion " ability of the kidney. One-tenth of a gram of this dye in a capsule 
is given by mouth Or 0.05 gm. intramuscularly (ice. ofai :2o solution). 

The elimination of methylene blue, first as a colorless chromogen, begins 
in from 15 to 30 minutes after a subcutaneous injection. In from 3 to 5 
minutes later the elimination of the greenish-blue pigment begins. For it 
to appear only after an hour is pathological. Normally the excretion of 
this dye reaches its maximum in from 3 to 4 hours and continues from 
35 to 50 hours (48 to 60). About one-half is eliminated in the first 24 
hours. From the first the test has been severely criticised. In some cases 
the dye is entirely destroyed in the body and under normal conditions 
only about 50% of the amount injected is excreted through the kidneys. 

Indigo-carmine Test (Voelcker and Joseph). — Twenty cubic centi- 
meters of an 0.4% solution of indigo-carmine is injected into the gluteal 
muscles. The color of the urine of a normal person will change to a greenish- 
blue in from 10 to 15 minutes after the injection and return to normal in 
about 1 2 hours. (That is, its elimination is more rapid than that of methy- 
lene blue.) This test is not satisfactory since only about 25% of this dye 
injected is excreted through the urine and the rest cannot be accounted for. 186 

Rosaniline (Lepine). — One cubic centimeter of a 1% solution of 
rosaniline is injected into the gluteal region. The excretion of the dye 
begins in about 30 minutes, reaches its maximum in 2 or 3 hours, and con- 
tinues from 20 to 24 hours. Since from 65 to 95% of this dye is excreted 
in the urine this test theoretically has much in its favor. 

Salicylic Acid Test. — This test was adapted to clinical use by Widal 
and Ravant as a measure of renal permeability. The excretion of this 
acid is supposed to be through the glomeruli and to be governed by physical 
laws alone. One cubic centimeter of a 30% solution of sodium salicylate 
is injected (with a little cocaine to reduce the pain) intramuscularly. The 
urine is examined at the end of K hour, then hourly, by the addition of a 
10% Fe 2 Cl 6 solution for its appearance and then the amount eliminated is 
determined colorimetrically. 

Normally the violet color will appear at the end of half an hour. It may 
even in 15 minutes. It reaches a maximum in from 1 to 3 hours and dis- 

186 This test is, however, preferred to all others by Thomas and Birdsall, Jour. Amer. 
Med. Assn., 1917, lxix, p. 1747. 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 319 

appears in from 8 to 12 hours. The amount excreted (i.e., the per cent, of 
the total) in 5 hours is taken as standard. In the various forms of nephritis 
the excretion may begin within the first half hour and reach a maximum 
at the same time in all, but in some cases of interstitial nephritis the output 
is continued over a longer time. Striking exceptions, however, are reported. 
This has certain advantages over the similar potassium iodide test since 
it is simpler and more rapid. Zeigan 187 recommends the quantitative 
estimation of the salicylic acid output and Singer 188 that of the KI. 

The Phenolsulphonephthalein Test. — Rowntree and Geraghty 189 
proposed a test for the functional activity of the kidneys which has proven 
the best of all. The solution of phenolsulphonephthalein is made up 
as follows: 

Six-tenths of 1 gm. of phenolsulphonephthalein and 0.84 c.c. of 2 A 7 
NaOH solution are mixed with a 0.75% NaCl solution so that 1 c.c. of the 
solution will contain 6 mgms. of the dye. The result is the mono sodium, 
or acid, salt which is red in color and slightly irritating when injected. 
Two or 3 more drops of the 2 N hydroxide therefore are added which will 
change the color to a beautiful Bordeaux red. 

Twenty minutes to half an hour before the test the patient drinks from 
300 to 400 c.c. of water in order to insure a free urinary secretion, other- 
wise a delayed appearance of the dye may be due to lack of secretion. 

By means of a catheter the bladder is completely emptied. Then 

1 c.c. of the above described solution is injected intravenously in the upper 
arm by means of an accurately graduated syringe or injected intramus- 
cularly. The urine is allowed to drain into a test-tube in which has been 
placed a drop of 25% NaOH solution and the time of the appearance of 
the first faint pinkish tinge is noted. If the patient has no urinary obstruc- 
tion the catheter is withdrawn on the appearance of the drug in the urine 
and the patient is instructed to void at the end of 1 hour and again at the 
end of the second hour. A rough estimate of the time of appearance of the 
dye can be made without the use of the catheter if the patient will void at 
frequent intervals. In cases of prostatic trouble, however, it is wise to 
leave the catheter closed by a cork stopper in place until the end of the 

2 hours. The bladder is thoroughly drained at the end of the first hour 
and again at the end of the second. 

Each sample of urine is measured and its specific gravity recorded. 
Sufficient NaOH (25%) is now added to elicit a maximum brilliant purple- 
red color. The urine is now poured into a measuring flask and enough 
distilled water added to make its volume up exactly to 1 liter. It is then 
thoroughly mixed, and a small filtered portion used to compare in the 
Duboscq colorimeter with a standard solution, or to read against a color 

187 Centralbl. f. uni. Med., 1903. 

188 Zeits. f. klin. Med., 1903, xlviii, p. 157. 

189 Jour, of Pharm. and Exp. Therap., July, 1910, vol. i, No. 6. 



320 CLINICAL DIAGNOSIS 

scale in the Autenrieth-Konigsberger hemoglobinometer (sometimes called 
the ' ' Hellige ' ' after the maker) . This instrument as modified by Rowntree 
and Geraghty has been found easier to use and as satisfactory as the 
colorimeter. 

The standard solution used for comparison is made up by adding 
3 mgms. of phenolsulphonephthalein (or % c.c. of the solution injected) to 
i liter of water which has been made alkaline by the addition of i or 2 
drops of 25% NaOH solution. This beautiful purplish-red solution will 
retain its intensity of color for weeks. 

One cup of the colorimeter (right) is half filled with this standard solu- 
tion and the plunger lowered so that the indicator reads 10. Some of the 
diluted urine (depending on the intensity of the color) is placed in the other 
cup, and the plunger manipulated until the 2 halves of the fields have an 
identical intensity of color. The indicator of the left plunger is now accur- 
ately read, and the amount of dye estimated. If, for example, the urine 
side reads 20 and the standard 10, then the diluted urine must contain 
only Y 2 as much dye as the standard solution. To estimate the percentage 
of dye excreted in the urine one multiplies the reading of the standard by 
100 and divides by the reading for the solution containing the urine, e.g., 

=50 which indicated that there is 50% as much dye in the urine 

as in the standard solution used for comparison. This compares the amount 
of dye in the diluted urine with that in the standard used for comparison, 
but to estimate what percentage of the drug administered is excreted one 
must compare the amount excreted with 6 mgms. rather than 3 mgms. 
The result in the example given above is 50% of the 3 mgms. or 25% of the 
6 mgms. (the amount injected) so that the excretion is 25% of the amount 
administered. It is possible by this method to detect a difference of 0.04 
mgm. in the output of phenolsulphonephthalein. 

The standard for comparison described above was chosen arbitrarily, 
because of the beautiful pink color which such solution presents when the 
indicator stands at 10. The doses injected have varied from 3 to 60 mgms. 
but 6 mgms. was selected as most satisfactory in the majority of cases. 

The objection raised against the quantitative estimation of most dyes 
by colorimetric methods is that the normal coloring matter in the urine 
will interfere with an accurate estimation. In the case of phenolsulphone- 
phthalein this difficulty is slight since the color of this dye is so very 
brilliant. Even from 200 to 250 c.c. of urine may be added to the solution 
before it is diluted with distilled water to 1 liter without changing the read- 
ing. In the case of patients an excretion of more than 200 c.c. of urine an 
hour would indicate a polyuria and such urine would have a lower specific 
gravity and a paler color than normal. 

If the color of the urine is such that an error might arise from this source 
one can correct it by adding to the standard solution the same amount of 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 321 

a similar urine as is obtained from the patient. While in this way very 
accurate quantitative estimations can be made yet in the vast majority oi 
cases such correction is unnecessary since it is not often that a patient 
voids more than 250 c.c. in an hour. In the majority of cases the technic 
of the test is simplicity itself. 

In normal cases the dye appears in the urine in from 5 to 11 minutes, 
from 50 to 60% is excreted in the first and from 20 to 25% in the second 
hour; that is, from 60 to 80% is excreted in the first 2 hours. 

The excretion of the drug bears little if any relation to the excretion 
of water. A high output of dye has accompanied a small amount of urine 
and the quantity of dye excreted may be small when the amount of urine 
is great. It is immaterial, so far as the excretion of the dye is concerned, 
whether the urinary output during these 2 hours is 50, 200, 400 or 500 c.c. 

In acute nephritis the functional ability of the kidneys as determined 
by this test may fluctuate in 24 to 48 hours from one extreme to the opposite. 

In many cases of parenchymatous nephritis there is a marked decrease 
in the amount of dye excreted and this reduction would seem to run parallel 
to the amount of renal sclerosis present. In 1 case only 10% was excreted 
in 2 hours. In the early mild cases the function may be but slightly dis- 
turbed, while there is 1 group of cases where there is definite hyperperme- 
ability so that 80% and over may be eliminated in 2 hours. 190 . 

In chronic interstitial nephritis the output is low in practically every 
case and the decrease is proportionate to the degree of severity of the 
disease as estimated clinically. In several cases only a trace of the drug — 
less than 1% — was eliminated in the course of 2 hours. These patients 
nearly all die within 2 months, many, but not all, with uremic symptoms. 

In nephritis, contrary to the normal, the excretion of the dye during 
the second hour is usually greater than that of the first. 

In cases with obstruction in the lower urinary tract, e.g., hypertrophy 
of the prostate, this test is particularly valuable. These patients frequently 
have also nephritis, pyelonephritis, pyonephrosis, pressure atrophy, etc. 
The amount of their urine, its content of urea and of total solids may be 
practically normal and yet the patient be on the verge of a renal failure 
which will be precipitated by any operative interference. In these cases 
the phthalein test will differentiate those with severe from those with 
slight" renal damage. This test has demonstrated the greatest impairment 
of function in cases with large residual urine and who have not been leading 
a catheter life. Operations on these men have in the past often proved 
fatal and yet now may be safely performed after an adequate regime has 
resulted in a decided improvement of the kidney function, as indicated 
by this test. When the time of appearance of the dye is delayed beyond 
25 minutes and the output of drug for the first hour is below 20 %, operation 
is postponed, regardless of the patient's clinical condition. If under routine 

190 Baetjer, Arch, of Int. Med., June 15, 1913, vol. xi, p. 593. 
21 



322 CLINICAL DIAGNOSIS 

reatment the output remains low but constant the renal function is prob- 
ably in a stable condition and the operation may be cautiously performed. 
In such a case the operation would be postponed even though i test showed 
a large output of the dye, in order to determine whether or not this condi- 
tion is stable ; for it has long been recognized that the function of the kidney 
is extremely variable following an operation for the relief of retention. If 
the further tests indicated a decreasing function, operations should not be 
performed unless unavoidable, since in our series death from acute suppres- 
sion sometimes follows operation. 

Again, operation should not be attempted when but a trace of dye is 
excreted, since grave renal changes certainly exist. Two patients whose 
output of dye during 2 hours was but a trace died of uremia within a short 
period, though at the time of the first test no evidence of uremia was 
clinically present. In neither case was any operation performed. 

In no case in which this test indicated, prior to operation, an efficient 
or stable renal function has any evidence of renal insufficiency become 
apparent subsequent to operation. 

This test is particularly valuable in determining the functional value 
of the individual kidneys. The separated urines are obtained by ureteral 
catheters. 

In normal persons the time of the appearance of the drug from the 
2 sides is almost the same (from 5 to 10 minutes). The occasional delay 
of 1 side of 2 or 3 minutes has been noted. In 1 case it appeared in 6 
minutes on the left side, and in 25 minutes on the right. In this case, 
however, there was an anuria on the right side, probably reflex, but the 
collection of urine for 1 hour showed equal secretions of drug from the 2 
sides. In only two normal cases has a distinct difference in the amount 
of drug excreted been observed. 

Seventeen cases of unilateral or bilateral renal infection were studied 
by Rowntree and Geraghty. Many of these cases came to operation, 
thus allowing opportunity to estimate the true value of the test. 

Unilateral Cases. — When only one kidney is diseased the appearance 
of the drug on the diseased side is delayed and the amount excreted is 
relatively and absolutely decreased. The actual time is of comparatively 
little value since it is the quantity excreted during a period of at least 
1 hour which is important. It is possible by extending the observations 
over a period of 2 hours with the catheters in place to demonstrate with 
some degree of accuracy the functional ability of each kidney. 

In the majority of cases of unilateral disease although the most of 
the work is done by one yet the combined output is almost equal to 
that of 2 normal kidneys. Following nephrectomy the 1 kidney will 
eliminate that amount of drug which would normally be excreted by 2 
healthy kidneys and a little more than the combined output of the 2 
kidneys prior to operation. 



THE URINE: FUNCTIONAL RENAL DIAGNOSIS 323 

While for practically every functional test great claims are made at 
first, which later must be greatly discounted, this test alone has thus far 
stood searching criticism by many workers and is growing in popularity 
year by year. It is of more value in prognosis than any other single test. 
While it should not be used to the exclusion of all others, yet in most 
cases its results indicate with fair accuracy the functional efficiency of the 
kidney. This test may never do any positive harm and yet that it is not 
entirely innocuous is evident by the slight increase in pulse rate and the 
slight fever which follows often and which may last for from i to 3 days. 

Schlayer's Lactose Test. — Two and one-half grams of lactose dis- 
solved in 25 c.c. of freshly distilled water, in small Erlenmeyer flasks 
stoppered with cotton, are pasteurized for 4 hours for 4 successive days at 
75 to 8o° C. Twenty cubic centimeters of the contents of 1 flask (contain- 
ing therefore slightly over 2 gms. of lactose), the preparation of which has 
just been completed, is injected intravenously. Only very slight consti- 
tutional symptoms follow the injection (a slight headache, malaise, rarely 
a chill and fever for a few hours) . The normal time for the excretion of 
this amount of lactose is from 4 to 6 hours. The urine should be collected 
4 hours after the injection and then for each succeeding 2 hours till 12 
hours have passed. The presence of lactose, in the urine is determined by 
Nylander's solution (using for each test the same amount of urine and of 
reagent and boiling each the same length of time) and its amount estimated 
by the polariscope. 

The mechanism of the excretion of lactose seems to differ essentially 
from that of phthalein. 

Phlorizin Test. — This test of the " secretory ability " of the renal 
epithelium, rather than of its " permeability " (in the latter function 
osmosis is supposed to play the important part, in the former, none), was 
proposed by Achard to replace the hippuric acid test (the ability of the 
kidney to transform benzoic to hippuric acid — this was a theoretically 
good test of renal functional ability but clinically useless since the quanti- 
tative determination of hippuric acid is so inexact) . The phlorizin test 
is based on the generally accepted opinion that phlorizin diabetes is due 
to a specific excretory activity of the renal epithelium and that a diminished 
or absent glycosuria would mean disease of these cells. The bladder is 
emptied and 1 c.c. of a fresh 1 : 200 solution of phlorizin (hence 0.005 g 111 -) 
is injected subcutaneously (a small dose is chosen which will produce 
glycosuria in only normal kidneys). Sugar is tested for at 15 minute 
intervals. In normal persons it will appear in from % to 1 hour and continue 
for from 2 to 4 hours. From 0.5 to 2.5 gms. of glucose will be eliminated. 
In nephritis as a rule less than 0.5 gm. of sugar is eliminated and in some 
cases none. This test is now considered unreliable. Some normal persons 
at times do not react at all and the variations in other normal persons may 
be greater than those seen in cases of renal disease. The test does not 



324 CLINICAL DIAGNOSIS 

permit one to separate the various forms of nephritis, since the " hypogly- 
cosuria," and " aglycosuria " occur with about equal frequency in all 
forms. Yet it is a test of renal activity and therefore quite different from 
the other. 191 

The Value of These Tests. — In the medical wards these tests certainly 
add to the clinical pictures which patients present. By means of them we 
can follow the results of treatment and in masked renal cases we have no 
better means of arriving at a prognosis. In preparing patients for operation 
this test has great value. It is our rule not to transfer any patient for oper- 
ation with an output of the dye under 50% if the operation can be delayed. 
The surgeon finds this test of great value, when, e.g., the question is the 
removal of a diseased kidney. In such a case the first question is, Is there 
another kidney ? The second is, Can this second kidney do the work of both ? 

Surgeons often get results with this test which are more definitely 
positive than those obtained in the medical wards. One reason for this 
may be that surgeons deal more with conditions which destroy all renal 
function (abscess, cancer, etc.), while among the medical cases there are 
so many in which many of the functions which can be tested are normal, 
but that unknown but all-important one, failure of which means uremia. 
Kummel 192 stated that in a long experience (in over 500 cases) he had never 
been decewed by cryoscopy of the blood, while Casper and Richer 193 con- 
sider the cryoscopy of the separated urines, together with the phlorizin 
test (determination of the amount of sugar eliminated by each kidney) — 
neither test alone but the agreement of both — of actually greater value 
than the microscopic or gross examination of the renal tissue. We quote 
these authors merely to show that a skillful man may get great assistance 
from any test, even a poor one, if only he is thoroughly acquainted with it. 

Disturbed Renal Function in Conditions Without Renal Disease. — In 
certain pathological conditions without definite renal disease the renal 
functions as measured by dietary tests are similar to those found in patients 
with advanced chronic nephritis. Christian 194 found this true in cases of 
pernicious anemia and ascribes it to the anemia, either a nutritional or toxic 
disturbance of the renal cellular activity. Mosenthal found it true also 
of cases of prostatic hypertrophy, pyelonephritis and polycystic kidney. 

DISEASES OF THE KIDNEYS 

At this point it is desirable to define more accurately the elements which 
enter into our ideas of nephritis. As our nomenclature testifies, it has 
been the opinion of pathologists that in addition to acute renal inflamma- 
tions there is also possible an interstitial nephritis, meaning by this a pri- 
mary proliferation of the fixed tissue-cells of the renal stroma. As clini- 

191 Hugnat and Revilliod, Arch. gen. de. Med., 1902, vol. viii, p. 19. 

192 Centralbl. f. Chir., 1903, vol. xxx, II, p. no; also Arch. f. klin. Chir., Bd. 67. 

193 Functional Diagnosis of Kidney Disease, 1903. 

194 Arch, of Int. Med., 1916, vol. xviii, p. 429. 



THE URINE: DISEASES OF THE KIDNEYS 325 

cians responsible for the lives of our patients we would maintain that in 
nephritis there are but two processes to be considered ; an active destructive 
process to be fought and a healing process to be favored. We would main- 
tain that in the great majority of cases the kidney suffers only in the dis- 
charge of its duty and seldom is it the primary seat of a disease. There 
is no " vicious circle " which explains the progress of a nephritis. If a 
nephritis continues it is because a cause (usually chronic infection) else- 
where in the body is continuing and, this removed, no organ can show more 
brilliant proof of attempts to return to normal than does the kidney. The 
acute infections or intoxication which injure the kidney are for the most 
part fed through the blood stream. These injure or destroy renal epithe- 
lium and interstitial tissue and the increase in the fixed tissue elements 
is a conservating process. There are, therefore, 2 processes in progress 
in each case of nephritis whatever the type, the acute destructive process 
and the processes of repair. Sometimes the former is very slight but like 
a slow smouldering fire it may in years reduce the total amount of renal 
epithelium to very small volume. 

The albumin, casts and renal, pus and blood-cells in the urine are evi- 
dence of this acute process. From the rapidity of elimination of various 
salts and nitrogenous bodies and from the chemistry of the blood-plasma 
we may judge of the total efficiency of the kidney. The more normal the 
kidney prior to an acute toxic injury the more spectacular will the result be. 
This is why the urine in an acute nephritis or following a foot-ball game 
may give a startling picture: intense albuminuria, casts of all descriptions, 
hematuria, etc., and yet the prognosis be very good, while the urine of a 
man dying in uremia may pass as " almost normal " since it contains 
" only a trace of albumin " and " an occasional cast." 

Albuminuria. — Since the presence of albumin in the urine would seem 
to be the most delicate test we have of renal irritation, we desired first to 
get a general idea of the incidence of albuminuria, the conditions in which 
it most commonly occurs and, if possible, to obtain some clue for further 
investigation. We 195 therefore abstracted the histories of 3631 hospital 
medical cases with satisfactory urine reports, taking them in order of 
admission to the hospital without reference to their diagnosis. 

It soon became evident that these patients must first be grouped 
according to age before any classification according to disease was possible. 

The age epochs we chose were : from 1 to 1 5 years, 1 6 to 2 5 , 2 6 to 3 5 , etc. 
The reason for choosing these figures is that the ages of 1 5 and 2 5 are more 
truly transition points in a person's life than are 10 and 20. The sexes 
also should be studied separately for certain decades at least. On the 
whole, however, sex has less influence than one might expect. We next 
divided the cases into 3 groups — those in which the urine was albumin-free 
throughout their stay in the hospital, those in which the albumin was pres- 

195 Jour. A. M. A., Jan. 6 and 13, 1906. 



326 CLINICAL DIAGNOSIS 

ent for a time but disappeared while the patient was under treatment and 
those in which albumin was present at each examination. The patients 
diagnosed as " neurasthenics " probably form a group of hospital patients 
as nearly normal as any, for this very diagnosis then meant that careful 
(for that period, 1 890-1 904) examination had given no evidence of any 
definite disease. Of the males with this diagnosis the percentages with 
albumin-free urine were : 1 to 1 5 years, 1 00% ; 16 to 25, 87%; 26 to 35,99%; 
36 to 45, 90%; 46 to 55, 84%; 56 to 65, 70%; and from 66 years and over, 
66%. The drop at the period of adolescence is interesting (see page 227). 
Of course no one would claim that these patients were normal. Doubtless 
now with improved methods of examination a positive diagnosis might be 
easily made in similar cases. Nevertheless this curve of incidence may be 
used as a base line in judging of the effects of the known disease. 

Of the fevers, typhoid after the twenty-fifth year is accompanied by a 
transitory albuminuria (febrile) in 30% of the cases and a persistent 
albuminuria in about 30%. One would expect higher figures than {his 
since the fever is so long-continued and bacilluria so common (about % of 
all cases). Yet as a disease of the past history, typhoid fever, strangely 
enough, seems to have injured the kidneys least, notwithstanding the 
deleterious influence which it has on the peripheral blood-vessels. 

Malaria of the tertian and quartan types has little effect on the kidney 
but estivo-autumnal much. Pneumonia has the highest percentage of 
transitory albuminuria of all the fevers we studied (in but about 25% of 
the cases was the urine albumin-free), but it had almost no permanent 
effect. Pulmonary tuberculosis and acute articular rheumatism cause but 
little febrile albuminuria. Of the afebrile diseases, the neurasthenics are 
the best off and those with arteriosclerosis, the worst. In fact, arteriosclero- 
sis seems the one dominating element among the causes of albuminuria. 

In those cases which came to autopsy a comparison was made between 
the anatomical lesions and the urinary findings during life. Cases with 
marked cloudy swelling, but no other renal lesion of autopsy, had had, as 
a rule, an albuminuria, usually slight, for 2 or 3 weeks before death. In a 
few cases the urine was albumin-free even shortly before death. Casts had 
accompanied the albumin, usually hyaline, but also waxy, epithelial and 
blood-casts. 

Fatty kidneys (no other microscopical changes) had developed in vari- 
ous diseases. Those with fatty infiltration were found in diabetes mellitus, 
pregnancy, etc. ; those with fatty degeneration in cases with various poisons. 
The amount of urine had been normal in most of these cases although in 
some severe ones it was decreased in amount; albumin from a trace to a 
a large amount had been present in every case, but in none for over 2 
weeks while casts had been present in a relatively large number of cases: 
hyaline, granular, fatty, and epithelial. The red corpuscles were few or 
many in number. 



THE URINE: DISEASES OF THE KIDNEYS 327 

The urine of patients whose kidneys at autopsy showed only chronic 
passive congestion was at first scanty in amount, dark in color and very 
acid. Its specific gravity varied between 1.025 an d 1.030. The urate 
sediment was often abundant. Urobilin and uroerythrin were increased 
and sometimes bilirubin was present. Sooner or later albumin appeared 
in traces, later in larger amounts, i.e., 0.1% while in 1 case it was 0.6%. 
Casts were present, chiefly hyalines, rarely the granular, yet on some 
days the hyaline, granular, waxy, epithelial and fatty casts were 
present in large numbers. A very few leucocytes also were found and 
still fewer red cells. The points of importance in the urine of this 
condition are: the small amount of albumin, the large urate sediment, 
the absence of renal epithelium and the scarcity of granular casts and 
leucocytes. A diagnosis of nephritis had been made clinically in over half 
of these cases. 

Acute Nephritis. — The pathologist studying the kidneys describes 10 or 
more forms of acute nephritis but the clinician finds classification very 
difficult. Senator separated tubular or acute parenchymatous from an 
acute diffuse nephritis, not as 2 distinct diseases, but as the extremes of a 
series of cases which includes every transitional form. In cases of the 
acute parenchymatous nephritis the tubules especially would seem to be 
involved; the glomeruli little, or not at all. The clinical symptoms, if any, 
are slight. The urine is diminished in amount, has a rather high specific 
gravity, contains but a trace of albumin and few or no casts. A heavy 
sediment often settles which consists chiefly of renal epithelial cells (hence 
the name ' ' nephritis desquamativa ") . These may occur singly or in casts. 
In the middle of the series are the cases with hyaline casts, sometimes few, 
sometimes many, crystals of uric acid and calcium oxalate, red blood- 
corpuscles, hemoglobin in granular casts or masses and a few leucocytes 
in the sediment. In some the small amount of albumin presents a remark- 
able contrast to the large amount of sediment and may be chiefly Morner's 
body. At the other extreme of this series are those of acute diffuse 
nephritis, a good illustration of which is that following scarlet fever. In 
this clinical symptoms are much more severe. The urine is diminished 
in amount; there may, indeed, be anuria for the first 24 hours. Other 
cases void from 50 to 100 c.c. for the first day or so and later from 200 to 
500 c.c. Toward death the amount may be diminished or increased. The 
specific gravity is normal as a rule, from 1.015 to 1.017, but in some cases 
is high, from 1.023 to 1.025 (when the amount of urine is from 300 to 600 
c.c.) while in the cases with very scanty output (under 500 c.c. in 24 hours) 
it may reach '1.030. The urine is usually of a dark color and cloudy, but 
in very mild cases it may appear normal. Blood is practically always 
present, in traces or enough to impart to the urine a slight smoky tinge. 
Some urines are reddish brown, brownish or even of a chocolate color 
depending on the amount of blood present and on the proportion of the 



328 CLINICAL DIAGNOSIS 

hemoglobin which has been transformed to methemoglobin. The albumin- 
uria is usually intense and yet in some cases, even fatal ones, mere traces 
may be present and these but for a few days, alternating even till death 
with periods during which the urine is albumin-free. Serum albumin and 
serum globulin both are present, and if many cells are in the sediment a 
certain amount of true nucleo-albumin and of albumose. Albumose in 
some cases is the only proteid found. Why, is not clear. This may explain, 
however, cases described as " albumin-free," since the examiner may have 
used only the heat and acid test, which would not precipitate albumose. 
As a rule not above i% of albumin is present and of this considerable is 
globulin. Red blood-cells may always be found in the sediment, also mono- 
nuclear cells, a few polynuclear leucocytes and epithelial cells from the urin- 
ary tubules which may be single or in masses and which usually are very 
fatty. Among the crystals met with are uric acid and calcium oxalate. 
Hemoglobin may be present either in amorphous granules or in casts. 
The leucocytes were very abundant in i case of acute nephritis with mul- 
tiple abscesses. The number of casts varies much from day to day. Some- 
times they are present in enormous numbers and in all forms. The epi- 
thelial, hyaline and coarsely granular casts predominate, but blood and 
leucocyte casts may also be present. As a rule the number of casts runs 
roughly parallel to the amount of albumin. In i case of acute hemorrhagic 
nephritis with areas of complete necrosis the amount of albumin just 
before death was slight but the number of casts, including leucocyte and 
granulars, was large. In i case of general septicemia traces of albumin 
were present in the urine on some days, none on others; and yet this urine 
often contained blood cells and hyaline and leucocyte casts. 

During the course of a case of nephritis the urine shows every symptom 
of renal insufficiency. The nitrogen output, apart from variations due to 
the diet, is diminished; the output of chlorides and phosphates is low hence 
the molecular concentration of the urine is less than normal. The uric 
acid output is about normal while that of the xanthin bases is said to be 
increased. The ability of the kidney to form hippuric acid is diminished 
and the glycosuria after phlorizin injection is either slight or absent. The 
phenolsulphonephthalein test gives varying results but in general gives a 
fair idea of the severity of the disease at the time the test was made but 
little as to prognosis. The renal test day may be quite misleading. The 
blood-urea estimations give the most consistently valuable means of deter- 
mining the degree of the progress in any given case. 196 In mild cases, and 
in severe ones as they improve, the urine is nearly normal. It is said that 
in acute renal infection the albumin disappears last but we believe the 
casts are more often found later than is albumin. 

Nephritis Hemoglobinuria. — In acute nephritis the urine may contain 
considerable hemoglobin and few or no red blood- cells. In certain cas es 

196 Atchlay, Arch, of Int. Med., Sept., 1918, xxii, p. 370. 



THE URINE: DISEASES OF THE KIDNEYS 329 

a hemoglobinuria would seem to be the cause of the nephritis, in others it is 
a symptom. The former may be true of cases of hemolysis due to poisons, 
burns, etc., while severe cases of the infectious diseases, especially typhoid 
fever, scarlet fever, malaria, Winckel's disease of the new-born, etc., may 
cause a nephritis with hemoglobinuria or nephritis and hemoglobinuria. 
Nephritis hemoglobinuria differs from pure hemoglobinuria in that in the 
former the amount of albumin is greater and the sediment richer in casts, 
renal epithelial cells, leucocytes and uric acid crystals. 

Acute Nephritis of Cholera. — With Asiatic cholera is said to develop 
a peculiar type of pure parenchymatous nephritis of the tubular variety. 
The urine is diminished in amount, in fact there may be anuria for from 
5 to 7 days. It is dark and cloudy, but rarely bloody, very rich in salts 
and may deposit a large urate sediment. Albumin is present in relatively 
larger amounts than in the other forms of parenchymatous nephritis. 
Hyaline and granular casts, renal epithelium, red blood-cells, leucocytes, 
uric acid and calcium oxalate crystals are found in the sediment. The 
urine is characterized also by its richness in the ethereal sulphates, the 
frequent presence of acetic acid and the increased amount of ammonia. 
The acidosis in these cases may be severe. One patient recovered after an 
anuria of 15 days. The condition of the urine improves much during the 
stage of reaction. 

Nephritis syphilitica acuta precox is sometimes marked by an intense 
albuminuria. In 1 case (Hoffman and Salkowski) the urine contained 
8.5% of albumin and coagulated to a solid mass when boiled. This urine 
had very little sediment, only a few casts, leucocytes and blood-cells. 

Subacute Nephritis; Chronic Parenchymatous Nephritis; Chronic 
Diffuse Non-indurative Nephritis; Large White Kidney. — This form of 
subacute nephritis, which may follow an acute nephritis or develop insidi- 
ously, is characterized clinically by its subacute course (it is usually fatal 
within 2 years) by the extreme anasarca and effusions into all the serous 
sacs, its incidence especially in young persons who work hard amid exposed, 
unhygienic surroundings and its frequent association with certain consti- 
tutional diseases, as tuberculosis, lues and malaria and with chronic 
alcoholism. 

The amount of urine during the acute stages of this disease is diminished 
to about 250 to 500 c.c. in 24 hours, the diminution varying as the edema. 
This is especially marked just before death. As the case improves, however, 
the amount increases, and if the patient be encouraged to drink fluids he 
may void from 5 to 6 liters of a very dilute urine each day. The amount 
is increased also when the edema or the effusions begin to absorb. Its 
specific gravity, which varies inversely as the amount, is, as a rule, almost 
normal or slightly increased, in some cases reaching even 1.040. Its reac- 
tion is faintly acid, but in some cases it is alkaline when voided and in all 
cases it quickly becomes so on standing. This makes the search for casts 



330 CLINICAL DIAGNOSIS 

difficult. Its color varies from a pale greenish-yellow to a red or a reddish 
brown. It is cloudy as a rule from the large amount of sediment present 
and foams easily on shaking because of the considerable amount of albumin 
it contains. Profuse painless hematuria may be a feature of this form of 
nephritis and unfortunately so far as diagnosis is concerned may be uni- 
lateral, first from i side then from the other. 197 

In this form of nephritis the albumin in the urine is abundant both 
relatively and absolutely. It varies roughly as the specific gravity and 
seems to bear no relation to the amount of edema present. It seldom 
reaches i% and for months may vary from 0.4 to 0.8%. In certain cases, 
however, it reaches 2% and Bartels reported one in which it varied from 
4 to 6%. The albumin quotient varies much. Nucleo-albumin is present 
in small amounts, also albumose. Some of these cases develop into the 
chronic indurative type in which cases the amount of albumin diminishes 
progressively. 

The output of urea is somewhat diminished even when there is much 
dropsy. That of uric acid varies, but remains within normal limits. The 
ammonia is normal. There is a certain retention of chlorine and of phos- 
phoric acid. 

The urine sediments in this condition resemble those of acute nephritis, 
but one finds more coarsely granular, fatty and waxy casts. Red blood- 
cells are always to be found and during the acute exacerbations many. 
There is little difference between the urine of the white and of the mottled 
kidneys except, perhaps, that in the latter are found more red blood- 
corpuscles, leucocytes and fatty cells. 

These kidneys show some functional insufficiency and yet in even the 
severe cases they do their work fairly well. The explanation for this is 
that the disease attacks locally successive parts of the kidney and that 
while 1 part is inflamed other parts can carry on the renal work. 

One would expect that an involvement predominantly glomerular 
would produce an intense albuminuria while one predominantly tubular a 
marked cylindruria. While in general this may be true clinically it is of 
little importance since both tissues are always involved. 

Except in the case of very young persons with a past history of fine 
health the diagnosis of subacute parenchymatous nephritis has difficulties 
since patients with an acute exacerbation of a latent chronic nephritis 
may present similar clinical features and may void a very similar urine 
and yet at autopsy small contracted kidneys be found. 

Chronic Indurative Nephritis. — A clinical subdivision of the group 
of cases which at autopsy have small contracted kidneys is exceedingly 
difficult. In fact even at autopsy the size and color of the kidney, and not 
the histological pictures, are the only safe basis of classification. Also, 
the anatomical conditions of the kidney may be of little help in interpreting 

197 Kretschmer, 111. Med. Jour., Aug., 1912. 



THE URINE: DISEASES OF THE KIDNEYS 331 

the preceding clinical course of the case since the same end result may be 
reached by different pathological processes (Christian) . 

Some have considered the so-called " senile atrophy," as almost physi- 
ological in elderly persons, claiming that the kidney grows slightly sclerotic 
with age. Nascher 198 for example says that " senile contracted kidney 
with slightly diminished output of urine of rather high specific gravity 
and a trace of albumin without casts is a physiological condition. It 
requires no treatment." Others however believe that these changes are 
common not because these patients are merely beyond middle life but 
because they have for more years harbored infected noses, bad tonsils, 
and infected mouths (pyorrhea alveolaris) than in the case of younger 
persons. If the process in elderly persons were merely an atrophy there 
should be few qualitative urinary changes. But as a result of hard work 
(possibly), of chronic infections, especially those of the nose and mouth 
and colon, of various diseases, especially gout and lues and of certain 
poisons, as lead and alcohol, insidious slow inflammatory and degeneration 
processes develop. The result of these is inflammation and degeneration 
of the epithelial elements and this may be general or focal, subcortical or 
periglomerular and with a subsequent compensatory new growth of connec- 
tive tissue. The kidney becomes hard and firm, it shrinks in size and finally 
is but a remnant of an organ. Such kidneys show that a nephritis can 
heal, for we found at autopsy markedly contracted kidneys which could not 
possibly have been suspected from the urine voided before death. It is 
evident from the history that some of these cases are the result of a pre- 
ceding acute or subacute nephritis. Other cases die with all the symptoms 
of an acute nephritis and we to our surprise find at autopsy evidence of a 
marked nephritis of years duration of which the final illness was an acute 
exacerbation. 

Even the pathologist cannot find any definite basis of classification of 
these contracted kidneys except their weight, the thickness of the cortex 
and their color, and so have divided them into the " red " and " white " 
kidneys. In the red kidneys the arterial changes are a prominent feature. 
They are firm and beefy, the disappearance of the epithelial elements is 
extensive and the amount of fibrous .tissue considerable. Yet these kidneys 
are seldom as small as are the white. The latter kidneys are very small, 
are pale yellow in color and conspicuously fatty. Little cysts are numerous 
in the cortex. The student should be reminded that a considerable sclerosis 
of the smaller blood-vessels of the kidney, extensive enough to produce 
malnutrition of the renal epithelium, will lead to atrophy and sclerosis of 
the kidney; and that while this process is not nephritis yet it explains 
many of the lesions of nephritis. In each kidney there are 2 reasons for the 
connective tissue proliferation: the death of epithelium as the result of 
direct toxic action and the starvation of epithelium as the res ult of vascular 

108 N. Y. Med. Jour., June 24, 1916. 



332 CLINICAL DIAGNOSIS 

disease. The latter process may be in part local and i kidney therefore 
suffer much more than the other. Christian 199 after years of study finds 
himself justified in making the diagnosis " chronic interstitial nephritis " 
and " chronic glomerular nephritis " less and less often and now more 
often makes the diagnosis " chronic nephritis with or without hypertension." 

Chronic Interstitial Nephritis. — In chronic interstitial nephritis the 
renal tissues have suffered from a slow chronic infection which has gradually 
reduced the amount of secreting tissue until but very little may be left. 
These are the cases in which the acute element is slight but since it is in 
operation for years it destroys an immense amount of kidney tissue. These 
cases are marked by their very insidious onset. The only symptom for 
years may be a slight albuminuria with perhaps a few casts and these may 
be absent for long periods of time. The amount of urine is increased slightly 
at first, but later in a well-developed case from 2 to 3 liters and sometimes 
even 12 liters are voided daily. On the other hand it may at times sink 
to normal or even under. The urine is pale, clear, definitely acid and has 
a specific gravity constantly between 1.010 and 1.005 irrespective of how 
much water the patient drinks. This fixation of specific gravity shown 
clinically by the low specific gravity of the morning urine is always signifi- 
cant of this condition. The molecular concentration is diminished. The 
amount of albumin seldom rises above 0.05%, and usually is much less. 
It is often absent in the morning voiding and may indeed be present only 
after a day of unusual exercise or an especially hearty meal or some unusual 
excitement. 

Hyaline casts can usually be found in the centrifugalized sediment. 
Red blood-cells are very common in the sediment. Sometimes there is 
definite hematuria. There is often sufficient desquamation of the epithe- 
lium cells of the urinary tract to produce a cloudy urine resembling that 
of cystitis. 

In the cases of arteriosclerotic kidneys the albuminuria appears late 
and is often intermittent. The most of the cases of so-called " contracted 
kidneys with albumin-free urine " belong here, and in these cases albumin 
is found even more constantly and in larger amount than in cases of the 
preceding group. During periods of improvement the casts often disappear 
first leaving a pure albuminuria while in the group to which the small white 
kidneys belong the albumin often disappears first. 

The output of nitrogen is practically always normal but the percentage 
of the various nitrogenous bodies may vary somewhat. In uremia, e.g., 
the ammonia may rise at the expense of the urea. Uric acid is low and the 
xanthin bases are increased. The tests for functional renal efficiency some- 
times indicate a pathological insufficiency but more often do not. The 
sediment is scanty and difficult to find. After a long search but 1 or 2 casts 
m ay be found in a centrifugalized sediment. These usually are hyalines 

199 Cleveland Med. Jour., April, 1917. 



THE URINE: DISEASES OF THE KIDNEYS 333 

although sometimes they are finely granular casts. Sometimes one finds 
a few renal epithelial cells and a few leucocytes, and more often than one 
would think a few red cells, especially after exertion. Uric acid and calcium 
oxalate crystals are common in the sediment. 

During acute exacerbations of a chronic nephritis the urine may closely 
resemble that of more acute nephropathies. 

For the diagnosis of chronic nephritis one should examine the morning 
and the evening urines separately and also that voided after severe exercise. 
The urine of these patients may resemble so closely that of other conditions 
(e.g., the convalescence of acute or subacute nephritis, waxy kidney, and 
the cyclic, or " physiological " albuminurias) that the clinical history and 
the physical examination of the patient are necessary for diagnosis. The 
tests for renal function are of great value in these cases in determining 
diagnosis, prognosis and treatment but must be interpreted in the light 
of the clinical findings. 200 

Amyloid degeneration is a condition which may be superimposed 
upon any form of nephritis, of which it really forms no part. When the 
kidney is merely waxy, the urine is said to be normal. In the majority of 
cases the condition accompanies a nephritis and could not be suspected 
from the urine alone, the examination of which would suggest, when con- 
centrated, chronic passive congestion; when dilute, small contracted kidney. 
The classical description of the urine of waxy kidney is that it is increased 
in amount, is pale, clear, faintly acid, of a low specific gravity, 1.005 to 
1.012, that it contains abundant albumin with relatively much globulin 
and very few casts. This picture of Traube, however, is rare. The albumin 
may occur in traces or fail and the casts may be numerous. The casts are 
often fatty. Renal epithelium is seldom seen and red blood-corpuscles 
are extremely rare. 

Uremia is the name given a syndrome usually associated with severe 
renal conditions and considered the highest expression of renal insufficiency, 
the most marked features of which are cerebral in origin; confusion, mania, 
gradually developing coma and often convulsions. The name was given 
by Bright who found in such cases the blood urea much increased and 
who supposed this the toxic substance involved. In chronic nephritis the 
body would seem to become tolerant to the renal insufficiency, for uremia 
is less common in chronic than in acute nephritis. The interesting experi- 
mental work of Folin, Karsner, and Denis, 201 emphasizes the importance of 
glomerular rather than tubular lesions in the production of that nitrogen 
retention which, it is supposed, explains uremia. 

But renal insufficiency alone, even though lethal, does not explain uremia. 
There can be no more complete renal insufficiency than that which follows 
the removal by operation of all the kidney tissue a patient has, as when the 

200 Christian, The Jour, of Urology, June, 1917, I, No. 3. 

201 Jour of Exp. Med., Dec. 1, 1912, vol. xvi, p. 789. 



334 CLINICAL DIAGNOSIS 

surgeon excises the diseased kidney of a patient whose other kidney had 
previously been destroyed, or removes a double kidney, or which follows 
bilateral calculus or bichloride poisoning. In these cases death may be 
delayed for from 10 to 14 days, during which time and even until the 
last hour the patient's mind is quite clear and he shows to the end 
none of the symptoms usually associated with uremia. Again, we see 
patients with chronic nephritis who, for even 3 weeks before death, voided 
a normal amount of urine which was almost normal as judged by the 
ordinary tests, but who eliminated practically no phenolsulphonephthalein 
in 2 hours and whose blood-creatinin was well over iomgms. per 100 c.c. 
of blood, and who were clear of mind until the last hour or so of life. This 
is good evidence that the retention of urinary constituents alone is not 
enough to explain the coma or convulsions of uremia. 

Foster 202 has clarified our ideas much by dividing the cases of severe 
renal insufficiency into 3 groups : (1) the retention type, the urinary poison- 
ing of Ascoli, in which practically all the constituents of urine are retained. 
This type follows the removal of all the functioning renal tissue, mercuric 
bichloride poisoning, impacted renal calculus, etc. In these cases there 
are no mental or nervous features, no convulsions and no gastro-intestinal 
symptoms until the very end. These cases present a pure type of urinary 
poisoning. A somewhat similar type is seen in cases of small contracted kid- 
ney with arterial hypertension, the symptoms beginning with the terminal 
cardiac decompensation. The picture develops more slowly than in the 
above cases and ends with asthenia, anorexia, a mild delirium and stupor. 
(2) The cerebral edema type. The second type is more often seen in cases 
of large white kidney (subacute parenchymatous nephritis) which renal 
disease leads to retention of only certain of the urinary constituents, especi- 
ally of the water and salts. In these cases the cerebrospinal fluid is under 
increased pressure, there is edema of the retina and at autopsy edema of 
the brain and meninges. These cases when they develop uremia have 
vomiting, headache, stupor, amaurosis due to retinal edema and finally 
coma, but very seldom convulsions. 203 (3) The toxic type or epileptiform 
uremia. In this, the classical type, there is in addition to the nitrogen 
retention the presence of some toxin which leads to convulsions. This is 
the uremia of most authors. 

Foster emphasizes the scarcity of pure types of these forms of uremia, 
the features of more than one usually being present. 

One group of 10 of our cases was interesting since it suggested that 
uremia may sometimes be followed by an apparent improvement 
in the patients' condition. In 8 of our cases of terminal uremia the 
albumin increased before death. In 1 case of uremia there was but 
a trace of albumin on the day before and on the day following the convul- 

202 Jour, of A. M. A., 1916, vol. 67, p. 927. 

203 See Trans, of the Assoc, of American Phys., 19 15. 



THE URINE: DISEASES OF THE KIDNEYS 335 

sion, but on the day of the convulsion the albuminuria and the cylindruria 
were intense. 

In eclampsia the urinary features are similar to those in epileptiform 
uremia. 

The temporary character and the extreme grade of the albuminuria one meets 
with in eclampsia are striking. In i case in the maternity ward the urine at 10 a.m., 
March 6, contained 0.653% albumin (gravimetrically determined). The woman was 
then in the first stage of labor and it was then that convulsions began. The urine between 
10 a.m. and 5 p.m. of that day contained 1.23%; between 5 p.m. and 9 p.m., 0.19%; at 
midnight, 0.075%; at 3 a.m., March 7, 0.025%; and for the rest of that day and later 
merely a trace. 

In another case the total albumin was 0.4678 gm. per 100 c.c, of which the globulin 
was 0.16 gm. per 100 c.c. (34%). In still another case the urine contained 18 gms. of 
albumin per liter, a multitude of casts and renal epithelium and yet at autopsy the 
kidneys presented no evidence of severe trouble. 

Unilateral Nephritis. — In the Johns Hopkins Hospital series of cases of 
nephritis followed to autopsy there was no case of strictly unilateral nephri- 
tis, but there were at least 30 cases with a considerable inequality in size 
of the 2 kidneys and in 3 the difference was marked. In these 3 cases the 
combined weights of the kidneys were 155, 190 and 205 gms. and the differ- 
ence in weight between the 2 organs respectively 45, 50 and 65 gms. In a 
very interesting case at operation Dr. Kelly found unilateral suppurative 
nephritis. Reisman and Muller 204 have discussed this subject at length. 
They say that acute unilateral inflammations differ from the commoner 
types of acute nephritis in that they are interstitial in character, are inflam- 
matory rather than degenerative in nature and have a marked tendency 
to abscess formation. These cases occur in connection with scarlet fever, 
erysipelas, osteomyelitis, endocarditis, pyemia, pyelitis, etc.; that is, as 
part of a general infection and are the result of bacterial invasion of the 
kidney. 

Renal Atrophy. — Renal atrophy may be due to insufficient blood-supply, 
to cachexia, the anemias and especially to advancing age, the so-called 
' ' senile atrophy. ' ' One sees microscopically no great increase in connective 
tissue. The urine is practically normal and without albumin. 

Congenital Cystic Kidney. — The urine in the very rare condition of 
congenital cystic kidney may be normal but more often resembles that of 
small contracted kidneys. Its amount is increased, its specific gravity low, 
a trace of albumin may or may not be present while usually the urine con- 
tains considerable blood. The contents of these cysts are not at all uniform, 
not even in those of the same kidney. In some the fluid is clear, watery 
and almost colorless, in others it is milky or colloidal; some cysts contain 
urea even in large amounts, also uric acid, while others contain none. In 
some have been found cholestrol crystals, colloid or proteid-like masses 
and rosette masses which resemble leucin. 

204 Arch, of Int. Med., June 15, 1913, vol. ii, p. 601. 



336 CLINICAL DIAGNOSIS 

Suppurative Nephritis. — The urine of cases of suppurative nephritis 
contains albumin in varying amounts and but few casts. In the sediment 
of i case there were a great many red blood-cells and leucocytes while in 
that of another there were very few leucocytes. When the pus-cells are 
numerous the urine will be alkaline. In a very few cases the urine has 
contained fragments of renal tissue. 

In cases of purulent nephritis the amount of pus in the urine may be 
disappointingly small since a kidney with an abscess which involves the 
whole organ may excrete no urine. Sometimes the kidneys are studded 
with renal abscesses and yet the Urine quite free of pus since none of 
these abscesses communicate directly with the tubules. 

Cancer of the Kidney. — Hematuria is often an early, even the first, 
symptom of cancer of the kidney provided this involves a pyramid. 
Hematuria was a feature in % of the Hopkins cases and the first symptom 
in ]i of them. The amount of blood voided may vary from a very slight 
trace to a fatal hemorrhage, which may be intermittent or of long duration; 
the blood may be fresh or decomposed, while clots even of large size may 
be voided. Otherwise the urine of these cases is practically normal. 

Tuberculosis of the Kidney. — Cases of general miliary tuberculosis 
usually have no urinary symptoms and those which may be present are not 
due to the tuberculosis alone. In tuberculosis of the pyramids with the 
formation of large caseous masses which may break down leaving a cavity, 
the so-called " renal phthisis," the urine is similar to that of pyelonephritis 
except that caseous material may be found in the urine. If the pelvis is 
not involved there may be no urinary changes. In very early cases of 
renal tuberculosis polyuria with or without albuminuria is often an inter- 
esting feature. Hematuria also may be the first symptom in such cases 
and was present in 8 of the 17 cases reported by Dr. Walker. 205 This early 
hematuria is very seldom a marked or serious feature and may last for 
months. It is present both day and night and bears no relation to the 
position of the patient, hence differs from that due to calculus. On the 
other hand it may be so severe as to be a serious feature. Blood-clots 
often appear in the urine. Pus was present in 15 of the 1 7 cases, sometimes 
a little, sometimes large amounts, depending on the position of the cavity. 
The sediment may contain (in 9 of the 1 7 cases) tissue detritus in masses 
about the size of a grain of sand in which are found tubercle bacilli and 
elastic tissue. Albumin was present in 16 and casts in 6 of this series. One 
should not be misled by the perfectly normal urine which may be excreted 
during the days on which no urine comes from the diseased side. 

In general it may be said that in all cases of hematuria and pyuria, 

especially if the urine is acid, tuberculosis of the kidney should be excluded. 

For diagnosis the tubercle bacilli themselves must be found. And yet 

since these bacilli can.be excreted through a " practically normal " kidney 

205 Johns Hopkins Hosp. Rep., vol. xii. 



THE URINE: DISEASES OF THE KIDNEYS 337 

tuberculosis of other organs must also be excluded. If a focus of this 
disease does not ulcerate into the pelvis of the kidney the entire organ may 
be destroyed before the condition is suspected. 

Infarction of the Kidney. — An intense albuminuria which begins sud- 
denly, which disappears soon and which is associated with no abnormal 
sediment strongly suggests renal infarction. One usually finds, however, 
evidence of a preceding nephritis. The sediment usually contains red 
blood-cells but a marked hematuria is rare. 

In cases of bilateral infarction there may be oliguria and even anuria. 

Pyelitis and Pyelonephritis. — Inflammation of the pelvis of the kidney 
may be due (i) to an infection ascending along the ureter, to a descending 
renal infection, or to an infection extending by contiguity from neighboring 
organs; (2) to local causes, as stone, cancer, tuberculosis, parasites (echino- 
coccus, amebse, etc.), trauma and floating kidney; or (3) to systemic causes, 
especially to the specific toxins of acute fevers, to medicines, etc. It is 
usually unilateral. 

The symptoms of pyelitis are usually masked by those of the disease 
of which this is a complication ; but even when the possibility of a pyelitis 
is realized there may be so few local symptoms which indicate it that 
ureteral catheterization is necessary. 

The urinary features in pyelitis will depend on its cause. Sometimes 
there is anuria (due to the reflex influence of the diseased over the sound 
kidney) while in chronic cases the amount of urine is sometimes even 
trebled. The urine contains but little albumin. It is cloudy from the 
presence of pus, blood and mucus, and faintly acid unless it has undergone 
ammoniacal decomposition. In the diphtheritic form of pyelitis, the urine 
will contain fibrin threads and even casts of the pelvis of the kidney. 

In the pyelitis of infancy due to Bacillus coli there may be no pus in 
the urine until a few days after the temperature has begun to rise. 206 

Microscopically the urine contains red blood-cells, mucous fibers, pus 
and various epithelial cells; uric acid and calcium oxalate crystals, also 
phosphate crystals if the urine is alkaline when voided; fibrin coagula, 
tissue constituents and other elements suggesting the cause of the trouble, 
as tissue fragments, tumor fragments or parasites. All forms of the epi- 
thelial cells of the transitional epithelium will be present. It is possible 
that a preponderance of cylindrical tailed cells may suggest the renal pelvis 
as the seat of the inflammation (see page 264 and Fig. 51, a). We have 
found many of these cells, often in tile-shaped clusters, in the urine of 
several cases of pyelitis, but in 1 very acute case of pyelitis with autopsy 
the urine contained none. There will be no casts or renal epithelial cells 
in case nephritis also is not present. 

A particularly important sign of pyelitis is the variation in amount and 
quality of the urine. The temporary obstruction of the diseased side will 

206 Thomson, Quart. Jour. Med., 1910, vol. iii, No. n. 
22 



338 CLINICAL DIAGNOSIS 

explain the periods with normal urine followed by periods with urine from 
the affected side. 

In the diagnosis of pyelitis the absence of disturbance of micturition 
is of great importance, also the homogeneous distribution of the pus in the 
urine and the club-shaped tailed cells in groups with a tile-like arrangement. 

In hydronephrosis, pyonephrosis and uronephrosis the urinary symp- 
toms of importance (apart from the pus) are the variations in the amount 
of urine, the periods of oliguria alternating with polyuria and the sediment 
the constituents of which will depend on the health of the cortex. 

Renal Calculus. — During an attack of renal colic the urine may be 
normal in amount or complete anuria may prevail. When, however, the 
obstruction is relieved, blood, mucus and pus will be voided with the urine. 

In addition to the colic, hematuria is a very common symptom of renal 
calculus (especially of the oxalate stones). Sometimes a clot of blood is 
passed, sometimes and especially early in the case, the hemorrhage is pro- 
fuse. Later the symptoms are those of pyelitis. 

With ureteral calculi are associated hematuria and oliguria, followed 
by po yuria. The oliguria was a feature in about 25% and anuria in about 
16% of Schenck's cases. 207 

Parasitic Diseases of the Kidney. — In echinococcus disease of the 
kidneys the only renal symptom may be a mucous catarrh of the pelvis, 
which later may become a purulent or gangrenous pyelitis. When a large 
hydatid cyst ruptures into the urinary tract there suddenly is voided a 
watery (or soapy, milky, or bloody) fluid. While the cyst is discharging 
the hooklets, scolices, fragments of membrane, etc., may be found in 
the sediment. 

For other parasites, see page 306. Pyelitis and even renal atrophy 
may be due to Bilharzia infections (page 306). 

207 Johns Hopkins Hosp. Rep., vol. x, p. 477. 



CHAPTER III 
THE STOMACH CONTENTS 

THE VOMITUS AND GASTRIC CONTENTS 

The various forms of vomiting have been grouped as follows: cere- 
bral, in brain and cord disease, as tabes, insular sclerosis, meningitis of 
brain or cord, cerebral anemia or hyperemia, concussion of the brain, 
brain tumors, etc. 

Toxic: opium, tobacco, ether, chloroform, alcohol, uremia, cholemia, 
pregnancy, etc. 

Periodic, " Cyclic," or " Recurrent " Vomiting- — These cases are 
characterized by the periodic recurrence of sudden attacks of vomiting, 
often without apparent cause, which are sometimes accompanied by inter- 
mittent hyper chlorhydria. There is evidence that some of these cases 
which resemble a secretory neurosis, especially those of children, are due 
to an acidosis, i.e., to an autointoxication. 1 

Neurasthenia and Hysteria. — One interesting case of neurasthenia 
vomited repeatedly from 3 to 4 ounces of bile-stained fluid in from 3 to 4 
hours after the stomach had been washed out. 

Reflex: as in peritonitis, strangulation of the bowel, sexual disturb- 
ances, chronic obliterative appendicitis, cholelithiasis, renal colic, intestinal 
worms, eye strain, etc. 

Local: due to gastric conditions, whether acute or chronic, and especi- 
ally those with stasis of the gastric contents. 

The Vomitus and General Considerations Concerning the Gastric 
Contents. — Considerable information, chiefly of a negative character, may 
sometimes be gained from the inspection of the vomitus. Its microscopical 
study seldom is valuable since we did not control the food ingested, while 
its chemical examination is often misleading for we seldom know the 
previous condition of the stomach, nor the character of this meal, nor can 
we control the time the food was in the stomach, nor evaluate the effect 
on digestion of the condition which led to the emesis, nor exclude the 
mucus and saliva from the mouth. 

The reaction of vomitus, with the exception of a few cases of achylia 
and of cancer of the stomach with alkaline gastric contents, is acid to litmus, 
provided the food had been in the stomach for at least half an hour, unless 
there had been a marked regurgitation of duodenal contents. This is of 
importance in excluding diverticula of the esophagus, in which case the 
vomitus may contain no gastric juice at all. Free hydrochloric acid is 
seldom present except in nervous cases since the conditions leading to the 
vomiting usually are those which would depress gastric secretion. 

1 Edsall and Snow, Am. Jour, of Med. Sci., 1904, vol. xxviii. 

339 



340 CLINICAL DIAGNOSIS 

The character of the vomitus is important. Abundant, thin, acid 
vomitus containing food eaten the previous day means dilated stomach 
due to pyloric obstruction ; very watery, thin, acid fluid with finely divided 
fragments of the food suggests ulcer; thick vomitus containing much mucus 
and pieces of poorly digested often decomposing meat suggests chronic 
gastritis and cancer of the stomach; recently eaten, undigested food sug- 
gests nervous vomiting. If the vomiting occurs at the height of digestion 
and during a paroxysm of pain which seems relieved by the vomiting one 
thinks of ulcer; if during or shortly after eating, of cancer, catarrh, or a 
neurosis; and if independently of eating (e.g., before breakfast) and con- 
tains not only mucus and bile but also food remnants, of gastric ectasis. 
Cerebral vomiting is often marked by a noticeable absence of effort ; morn- 
ing vomiting is suggestive of pregnancy, or, in the case of men, of alcohol- 
ism. In case of cancer or any other stenosis-producing disease at the cardiac 
orifice the vomiting immediately follows a meal; but if at the pyloric orifice, 
the vomiting occurs at least 3 hours after the meal and has a volume much 
greater than that of the meal. 

A small amount of blood in the vomitus is of no account since the effort 
of vomiting easily causes slight trauma of the esophagus or pharynx. 

Bile and Pancreatic Fluid. — Traces of bile are often present in the 
vomitus from a fasting stomach and in that raised with great effort. It 
has no significance unless it is constantly present and in vomitus expelled 
without strain sufficient to force bile from the duodenum into the stomach 
in which cases it might suggest stricture of the duodenum below the am- 
pulla. A green vomitus does not always contain bile, 2 for a " grass-green," 
" sea-green " or " dark-green " color may be due to the presence of algae 
or to chlorophyll-containing protophytes. One reason why vomitus from 
an empty stomach is more apt to contain bile than is that from a full one is 
that in the latter case the pylorus is more apt to be in tonic contraction. 
This may explain the former belief that in peritonitis the vomitus is more 
often bile-stained than in cerebral troubles, since in the former condition 
the stomach is often quite empty. 

Mucus is usually present in vomitus in large amounts. There are 
several reasons for this. One is that inflammations of the stomach wall 
are common causes of vomiting ; another, that while the amount of mucus 
secreted was normal the usual amount has not yet been digested; while a 
third is that the increased secretion of mucus represents a protective 
phenomenon, the wall thus shielding itself against chemical trauma, or 
from pus swallowed from the nose and mouth. This may explain why 
the vomitus of alcoholics usually contains large amounts of mucus. 

The vomiting of large amounts of acid gastric juice, sometimes 
pure, sometimes mixed with food, is common in cases of hypersecretion, 
especially in cases of gastroxynsis, a neurosis with periodic attacks of 

2 Kuhm, Zeitschr. f. inn. Med., 1902, No. 28; 1903, No. 1. 



THE VOMITUS AND GASTRIC CONTENTS 341 

vomiting of acid fluid. The proteid of the food in such vomitus will be 
well digested, the starch less so. 

The vomiting of large amounts of fluid containing food eaten 2 or 3 days 
previously, the proteid of which is poorly digested and the starch well 
digested, suggests malignant stricture of the pylorus while if the proteid 
is well digested and the carbohydrates poorly, this would suggest benign 
stricture, due to ulcer, etc. 

Fecal vomiting usually indicates a complete obstruction of the ileum 
or the colon, or paralysis of the intestinal wall due to peritonitis, etc. Such 
a patient vomits repeatedly, each vomitus a little more fecal than the 
former. That from the colon is black, foul-smelling and contains vast 
numbers of bacteria. Yet the absence of fecal vomiting would not exclude 
a total obstruction high in the jejunum and some hysterical patients have 
vomited the contents of the colon. For the vomitus to have even a sug- 
gestive fecal odor the obstruction must be at least 6 feet from the pylorus. 

Rice-water vomitus, seen in Asiatic cholera, is very watery in character 
and is filled with white flakes of mucous shreds and epithelial cells (see 
page 422). 

From the color and the odor of the vomitus a diagnosis of poisoning or 
alcoholism may be suspected. That of uremia has an ammoniacal odor. 

Some idea of the motility of the stomach may be obtained from an inspec- 
tion of the vomitus although temporary disturbance of gastric motility 
may be expected in many cases with conditions which would lead to vomit- 
ing. If food is vomited 7 hours or more after the last meal, gastric motility 
is certainly delayed, temporarily at least. At the end of 1 hour bread 
should have been broken up to a fine, crumbly sediment, which settles to 
the bottom of the glass. If the vomitus contains large particles of bread, 
and especially if these are coated with mucus, the secretion of hydrochloric 
acid is quite surely diminished. 

The chemical analysis of vomitus, as stated above, is exceedingly unsatis- 
factory. If free hydrochloric acid is present in vomitus, we may be sure 
that it is present in this case under more normal conditions; but if absent 
we can draw no conclusions. In general it may be said that normally both 
free hydrochloric acid and pepsin are present in the gastric juice 2 hours 
after a mixed meal. 

Tumor fragments have been found in vomitus but this is rare. In 
the vomitus may be found also round worms, segments of tapeworm, 
oxyuris, maggots, etc. 

Examination of the Fasting Stomach. — While the normal fasting 
stomach should theoretically be empty yet as a rule one can syphon off 
through a stomach tube from 1 to 50 c.c. of acid gastric juice. There is 
some difference of opinion concerning the amount which should be con- 
sidered the upper limit of normal for the fasting stomach, yet many agree 
with Boas that 100 c.c. or more of fluid certainly is abnormal and would 



342 CLINICAL DIAGNOSIS 

indicate hypersecretion or motor insufficiency. Which, may be decided by 
washing the stomach out at night and passing the tube the next morning. 
If the case were one of motor insufficiency the stomach will be quite empty. 
Riegel insists that the normal fasting stomach is always empty and that 
to find even a little fluid is pathological. 

The fluid from the fasting stomach is watery with a specific gravity 
from 1.004 to 1.005. It contains some free hydrochloric acid but not 
lactic acid and no bacteria. In many cases it is bile-stained, but this is 
not important unless it is found so on several examinations, in which case 
duodenal stricture may be suspected. If alkaline from the presence of 
pancreatic juice, trypsin may be tested for. To find trypsin in an acid or 
neutral fluid soda must be added at once to prevent the destruction of this 
ferment. Abnormal amounts of mucus may be present in cases of anacid- 
ity, atrophy of the mucous membrane, etc., but considerable washing is 
necessary to dislodge it. 

Test Meals. — The Ewald-Boas test breakfast consists of white 
bread, 30 to 40 gms., and water or tea without sugar or cream, about 
400 c.c. This should be taken at the time the patient is accustomed to 
breakfast. The bread should be masticated to a fine pulp and the whole 
ingested in not over 10 minutes. According to former methods the stomach 
tube was passed in just 1 hour from the completion of the meal and as much 
as possible of the gastric contents syphoned off. Normally one obtains 
from 30 to 70 c.c. of an acid fluid containing the partly digested bread in a 
granular condition and some mucus. If but little is obtained, as in cases 
of hypermotility, the meal was repeated on subsequent mornings varying 
the time before removal until confident that one is obtained while digestion 
is at its height. Unless from previous observation one is sure that the 
stomach empties itself well it should always be thoroughly washed out the 
night before a meal. The results of examination of the first test meal 
should be accepted with caution especially should they indicate disturbed 
motility or reduced acidity and the meal repeated until the patient has 
become accustomed to the procedure. 

Since bread is so variable in its composition, Dock substitutes for it 1 shredded 
wheat biscuit. 

Since the above breakfast consists so much of starch and water and contains so 
little proteid it cannot test well the most important of gastric digestive functions, that 
is, the hydrolysis of proteid. 

Riegel proposed a meal consisting of 1 plate of beef soup, from 150 to 200 gms. of 
beefsteak and 150 gms. of mashed potatoes. This should be eaten at the patient's 
regular dinner hour and removed at the end of from 3 to 4 hours. 

Fischer's meal consists of the ingredients of Ewald's test breakfast, plus a quarter 
of a pound of finely chopped, lean beef boiled and slightly seasoned. It is to be 
removed at the end of 3 hours. 

The study of gastric conditions by the chemical analysis of the stomach 
contents after a test meal has during the last 15 years fallen into almost 



THE VOMITUS AND GASTRIC CONTENTS 343 

complete disuse. The roentgenological examination is in part responsible 
for this. This will give certain data much more accurately than can the 
test meal and other data which the latter cannot give. And yet the ront- 
genological examination cannot replace entirely the chemical examination. 
One of the chief reasons why the latter has been so abandoned is that the 
profession has been careless in this work and so did not get convincing 
results. The introduction of the Rehfuss tube and the examination of 
specimens of the gastric contents removed each 15 or 20 minutes (see 
page 351) is a great advance in our methods and introduces a new era in 
gastric diagnosis. 

Among the common mistakes which have been made in gastric analysis 
are the following: 

The test breakfast should resemble as closely as possible the patient's customary 
meal. The Ewald breakfast is like the German early breakfast but does not at all 
resemble the American. It should be removed when digestion is at its height and this 
seldom is at the end of just 60 minutes. 

It should be eaten at about the same hour that that particular patient is accustomed 
to eat a meal of that general character. In this country the convenience of doctor and 
nurse rather than the habit of the patient is too much considered. The stomach should 
be thoroughly washed out the night before since a crumb of food of the day before can 
completely vitiate the results. 

The first meal is of practically no value, and often the second, because of the fear 
and disgust which the tube may inspire. 

Fischer has shown, and this probably illustrates the difference between any typical 
starch and proteid meals, that the results with his meal are much more constant than 
those with the Ewald breakfast. For instance, the diagnosis made early using the 
Ewald breakfast had to be changed after later meals in 40% of the cases, while in but 
about 8% with his meal. If he used both meals in the same cases the results were similar 
in 67% of the cases while 18% of those who showed hyperacidity with the Ewald break- 
fast showed less with his meal and 15% more. Of the cases subacid with the Ewald 
breakfast 30% were normal by his. In general it may be said that the Ewald breakfast 
will give some idea of what the stomach will do with an indifferent meal which excites 
but little secretion, while the Riegel will show the gastric functional ability when a 
greater tax is made upon the secretory cells. Some stomachs can handle the test break- 
fast well but not the larger meal, while others respond well only to the greater stimulus. 
Fischer gives several points of differential diagnosis based on the use of 2 meals. If 
the stomach be subacid to the breakfast but normal after the proteid meal we may 
suspect that the secretory structures are normal but that the constant presence of foods 
due to atony has reduced the sensitiveness of the mucosa ; if the gastric juice is subacid 
both to the proteid meal and to the breakfast, one may suspect organic changes; if sub- 
acid after the breakfast but hyperacid after a proteid meal, one might suspect defective 
innervation and the same would be true if it were hyperacid after the breakfast and 
normal after the largest meal. If the contents are hyperacid after the breakfast and 
still more so after the proteid meal one may suspect an increase of the oxyntic cells 
and especially so if the secretion continues for several hours after the meal. If the 
symptoms and an increased secretion both diminished after the meal, disturbed inner- 
vation may be suspected. Fischer emphasized the point that certain cases of 
dyspepsia, which for some time have been on an almost starvation diet, need not be 
fed up pretty well before a test meal is given. This may cause a gastric upset, but 
the flare-up of the condition will be an advantage in the diagnosis. 



344 CLINICAL DIAGNOSIS 

A point of importance in neurotic cases is that the time at which the 
meal is given should be chosen with reference to the symptoms, since at 
other times than during the nervous disturbances the gastric condition 
may be found normal. 

Acidity of the Gastric Juice. — The gastric contents to be examined 
should be tested first with litmus. This will indicate its reaction in general. 
If acid, this may be due to hydrochloric acid, free or bound, to organic acids 
and to acid -salts. In the great majority of cases the litmus will turn red; 
in a very few cases the fluid is alkaline. 

The fluid should next be tested for the presence of free hydrochloric acid. 

The gastric juice of the normal person is strongly acid from the presence 
of hydrochloric acid and certain acid-salts. This acid at the height of 
digestion after a mixed meal is present in 2 conditions : the bound and the 
free. By bound acid we mean acid which has entered into a loose combi- 
nation with certain organic bodies without disassociation of the acid- 
molecule and which in all ordinary chemical reactions, with the exception 
of certain color reactions, behaves as HC1. It will react acid to litmus 
and phenolphthalein but neutral to certain other delicate color reagents 
as Gunzburg's reagent, Congo-red, dimethylamidoazobenzol, etc. These 
acid-binding bodies in the stomach are proteins and many of its split 
products, especially the hexone bases, and mucus. 

There are in the gastric secretion also certain alkalies as sodium hydrox- 
ide and ammonia. These form with the acid neutral chlorides and take 
no further part in the gastric acidity. 

The normal stomach has a remarkable regulating mechanism which 
so controls its acidity that although there are great variations in the total 
amount of acid secreted, depending on the meal to be digested, the quali- 
tative relations of the fluid are quite constant for the same person at the 
same time after meals of similar composition. How remarkably accurate 
is this control is shown better by the constant differences which follow 
changes in the composition of the meals. 

The molecule of bound acid (which need not be hydrochloric) is neces- 
sary if pepsin is to split the molecule with which that acid is bound. Theo- 
retically an optimum digestion could be attained were there only just 
enough acid present to saturate all the acid-binding bodies, but the mucosa 
actually does maintain in the stomach during digestion a fairly constant 
excess of acid, that is, of hydrochloric acid, which is free since in excess of 
the acid-binding bodies. This free acid will give certain color reactions 
which the bound does not. This free HC1 probably has an important func- 
tion to play in sterilizing the gastric contents, in controlling gastric mobility 
and pyloric spasm and in the formation of hormones to stimulate the 
activity of other organs of digestion. 

In conditions of diminished secretion of hydrochloric acid the gastric 
juice may contain large amounts of organic (especially lactic) acid which 
also may aid the pepsin to split the protein molecule. 



THE VOMITUS AND GASTRIC CONTENTS 345 

The tests for free hydrochloric acid are for the most part color-tests 
for any free acid whether mineral or organic. 

Methyl violet is the indicator suggested by v. d. Velden, who first showed the pres- 
ence of free HC1 in the gastric juice. 3 This is still a very satisfactory reagent. One drop 
of the saturated aqueous or alcoholic solution of methyl violet is added to a test tube 
half full of water and then diluted with more water until it has a pale violet color. This 
fluid is then divided in 2 test-tubes; to the one is added the filtered gastric juice, to the 
other the same amount of water. Free HC1 will turn the violet to a fine blue color. 
This indicates 0.025% of free mineral acid; to produce the same reaction would require 
a much larger amount of free organic acid. 

Tropeolin 00 has been used, but is less sensitive than the above, indicating as it 
does but 0.03% of free HC1. The test is made in the same way as the above; free HC1 
will turn the yellow to a reddish-yellow color. 

The test easiest to use is Congo-red paper which a free mineral acid will 
turn to a sharp blue color, while free organic acids, even in strong concen- 
tration, will give a much less definite shade of blue. Acid salts, if strong, 
would give a positive test but not in the concentration found in the stomach. 

Dimethylamidoazobenzol is the reagent now most commonly used. In 
the presence of free mineral acid the yellow color of its solution changes 
to a fine pink. It reacts also, however, tc organic acids and acid phosphates 
in concentrations which might occur in the stomach. 

Gunzburg's solution (phloroglucin, 2 ; vanillin, 1 ; alcohol, 30) is the best 
test since it reacts only to free mineral acids and in gastric juice, therefore, 
only to free hydrochloric acid. One or 2 drops of this solution (which 
should be kept in a tightly corked blue bottle and not allowed to get too 
old) are gently warmed on a porcelain dish until just dry. One drop of the 
gastric juice is then allowed to come into contact with this and the gentle 
warming continued. If free acid is present a beautiful crimson line will 
appear at the edge of contact. This is a fairly sensitive test, less sensitive 
than the others, but of more value since it is final. 

It is to be emphasized that the above color-tests all indicate free acid, 
the most of them free mineral acid, and so free hydrochloric acid; that is, 
acid in excess of all acid binding bodies such as proteids, hexone bases, etc. 
Enough acid will have been secreted so that some will be free after a carbo- 
hydrate meal in from % to % of an hour; after meat, in from 1 to 1% hours; 
and after milk and potatoes, in % of an hour. 

Total Acidity. — The estimation of the total acidity of the stomach 
contents is the starting-point in all gastric analysis. This figure represents 
the amount of hydrochloric acid which would be present were all the acidity 
in the stomach at that particular time due to this acid; this, compared 
with the amount of free acid, gives a good picture of the secretory and the 
motor ability of the stomach. The total acidity is the sum of the free and 
the bound hydrochloric acid, of other acids, as lactic, butyric, etc., and of 
all acid-salts, e.g. (phosphates) present at that time. 

3 Deutsch. Arch. f. klin. Med., Bd. 23. 



348 CLINICAL DIAGNOSIS 

To 10 c.c. of gastric juice is added an indicator sensitive to all acid 
reacting substances. Tenth-normal NaOH is then added slowly from a 
buret, stirring all the time until the change of color shows throughout 
the whole volume of fluid. This titration may be done in a porcelain dish, 
a beaker, or an Erlenmeyer flask, the latter 2 against a white background. 

The indicator usually used is phenolphthalein, 2 or 3 drops of a 0.5% 
alcoholic solution. Among the others used are litmus, cochineal, methyl 
orange, etc. Phenolphthalein is preferred because of the sharpness of its 
end reaction. It is colorless in acid and brilliant red in alkaline solution. 
Yet it is perhaps the most unfortunate choice of all since it is not accurate 
in the presence of ammonia salts of which there is a fairly large amount 
in the gastric contents. The results of the titration will therefore be too 
high. The reason for its continued use is the desire to get comparable 
results to which an empirical value may be given. 

The unfiltered gastric juice, shaken well to a homogeneous suspension, 
should be used since the solid particles contain relatively more of the acid 
than do the fluid portions. 

The acidity of the gastric contents may be expressed in 2 different ways. 
If the number of cubic centimeters of 0.1N NaOH used be multiplied by 
0.00365 gm. we would have the weight of acid measured as hydrochloric 
acid neutralized, yet it is never the case that all the acidity is due to HC1. 

Table of Equivalents 
Acidity Gravimetric 
per cent. per cent. 

10 0.0365 

14 °-°5 

20 0.073 

27 0.1 

34 0.125 

40 0.146 

48 0.175 

50 0.182 

55 0.2 

61 0.225 

70 0.25 

73 °- 2 75 

80 0.292 

87 0.317 

90 0.329 

95 °-347 

100 0.365 

105 0.383 

109 0.4 

The better and usual method is to follow the suggestion of Jaworski 
who introduced the term acidity per cent, (abbreviation of this, A. P.) for 
the number of cubic centimeters of the alkali which would be required to 
neutralize 100 c.c. of the gastric juice. Since 10 c.c. of stomach contents 



THE VOMITUS AND GASTRIC CONTENTS 347 

is the amount used the titration figure multiplied by 10 will give the acidity 
per cent, without reference to the character of acid bodies which may 
be present. 

An illustration: if, using phenolphthalein as indicator, 10 c.c. of the 
gastric contents required 8 c.c. of 0.1N NaOH to neutralize the acids pres- 
ent, the acidity per cent, would be 80 A. P. Supposing that HO were the 
only acid present, then the gastric contents would contain 0.29% HC1. 
To avoid confusion, the symbol of gravimetric percentage is never used for 
" acidity per cent." 

Free Hydrochloric Acid, Mintz Method. — Ten cubic centimeters 
of the gastric contents are titrated with 0.1N NaOH until the test for 
free acid is no longer positive. This method assumes that the NaOH will 
neutralize the free before the bound HC1. 

Of indicators used undoubtedly the most accurate is Gunzburg's. As 
the sodium hydroxide is added, small drops of the stirred fluid are removed 
by a glass rod, or better still, a platinum oesa, and tested on a porcelain 
dish (see page 345). Fleiner would add 25 to 30 drops of the Giinzburg 
reagent directly to the gastric contents and then, as the sodium hydroxide 
is added, removes small drops which he warms in a porcelain spoon. Sahli, 
who also adds from 25 to 30 drops directly to the fluid, recommends 
that the glass rods with which the alkali is mixed with the gastric contents 
themselves be warmed, for the crimson color can be seen on the rod. Since 
a certain amount of gastric juice is lost in each of the Giinzburg tests the 
results should be confirmed using a fresh portion from which fewer drops 
need be removed. 

A much easier method, and one used in many clinics, is to touch the 
stirring rod after the addition of each fresh installment of alkali to a strip 
of Congo-red paper and to add the alkali until this no longer turns blue. 
Some to save time find the end reaction approximately with Congo-red, 
and then more accurately with Gunzburg's reagent. The easiest and most 
popular method of all employs as indicator a very small drop, the little 
which clings to the end of a glass rod, of dimethylamidoazobenzol. This 
in the presence of free acid takes a bright red color. The sodium hydroxide 
is added until the red just disappears. A drop of phenolphthalein is then 
added and the titration continued to determine the total acidity. 

The results with these 3 indicators are by no means the same. Those 
with Gunzburg's will always be the lowest and those with dimethylamido- 
azobenzol usually the highest. Those with Congo-red paper vary, but will 
stand between those of the other 2. 

Topfer's Method. — This method now is chiefly historical. The indicators used in 
the titration are dimethylamidoazobenzol for the free hydrochloric acid and then a 1% 
aqueous solution of alizarin for the bound hydrochloric acid. The latter indicator, 
however, has been abandoned, and the most accurate way of estimating the bound acid 
is to subtract the free from the total acidity. 



348 CLINICAL DIAGNOSIS 

Hydrochloric Acid Deficit. — In cases with an insufficient gastric 
secretion of HC1 it may be desirable to determine the acid deficit; that is, 
the amount of HC1 necessary to saturate the acid-binding bodies present 
in the gastric contents. This is done by adding 0.1N HC1 to 10 c.c. of the 
gastric contents until the test for free acid is positive. The amount neces- 
sary to add will depend on the amount of bound HC1 already present, the 
amount of proteid and bases which can bind the acid, and the amount of 
alkali secreted; hence a better term than HC1 deficit is that suggested by 
Sahli, "saturation deficit." Congo-red paper or Gimzburg's reagent can be 
used, but the former is sufficiently delicate. The determination of this 
value aids in following the progress of a case and in evaluating a therapy. 

Total Hydrochloric Acid. — The hydrochloric acid secreted by the gastric glands 
may be present in the gastric contents as free acid, bound acid, and as neutral chlorides. 
The total neutral chlorides includes those from the food, those formed in the stomach 
by the reaction of an alkali and the acid and those chlorides secreted as such. 

By " total hydrochloric acid " of gastric contents is understood the sum of the 
bound and free HC1 (" the physiologically active hydrochloric acid ") although some of 
the neutral chlorides were formed from the HC1 secreted as the acid. The Lutke-Martius 
method for determining this total HC1 is based on the principle that the difference 
between the total chlorine (a) and the chlorine left after incineration (b) represents the 
HC1 that was volatilized by heat. This method has been corrected by Reissner, who 
showed that NH 4 C1 also will be volatilized. He, therefore, first neutralizes the gastric 
juice with 0.1N NaOH using litmus as indicator. This neutralized fluid is then ashed 
and the chlorine determined a: 

0-£=HCl+NH 4 Cl, 
a-i = NH 4 Cl, 

d-b=nc\. 

The Arnold and Lutke methods are used in these determinations (see page 131). 

Determination of " a." — Ten cubic centimeters of the gastric fluid are measured 
with a pipet into a 100 c.c. measuring flask. Twenty cubic centimeters of Solution 
1 are added, the mixture stirred and allowed to stand for 10 minutes. A few drops of 
8% KMn04 are then added if necessary to decolorize the fluid. The flask is then filled 
with water to the 1 00 c . c . mark and the contents well mixed . This is then filtered through 
a dry filter until over % has passed through. Fifty cubic centimeters of this filtrate are 
measured into a beaker and Solution 2 then added from a buret until the resulting 
brown color is permanent. The number of cubic centimeters of Solution 2 necessary to 
precipitate the excess of silver are then multiplied by 2, since but half the filtrate was 
used in this titration. This product, subtracted from the amount of AgN0 3 originally 
added, will give the amount of AgNOs which was necessary to precipitate the chlorine. 

Determination of "b." — Ten cubic centimeters of the gastric juice are evaporated 
to dryness on a water-bath in a platinum dish. The residue is then burned over the 
free flame until the ash no longer burns with a luminous flame. It is not brought to a 
red heat, since this would volatilize some of the chlorides. The ash is then rubbed up 
well with water using a glass rod, extracted with about 100 c.c. of warm water, brought 
onto the filter and washed until a few drops of the filtrate no longer give a precipitate 
with AgN0 3 To the whole filtrate are then added 10 c.c. of Solution 1, and the deter- 
mination continued as for "a." 

Determination of "a." — Another 10 c.c. are first neutralized with 0.1N NaOH, 
using litmus as indicator, then ashed and the remaining chlorides determined as for "6." 

(a— b) X 0.0365 gm. =the per cent, of HC1. 



THE VOMITUS AND GASTRIC CONTENTS 349 

A bsolute A mount of Hydrochloric A cid Secreted. — The preceding methods are intended 
to give merely the percentage of the acid in the stomach contents at any one time. It 
is sometimes desirable to determine the total amount of acid present at a stated time. 
This method has some scientific, but no practical, value since considerable of the acid 
secreted will have already passed into the intestine. 

To determine this the stomach-tube is introduced and as much of the gastric juice 
as possible is syphoned out. Then 300 c.c. of water are allowed to flow in and out of the 
stomach several times. From the difference in the specific gravities of these 2 fractions 
the amount of gastric juice in the second fraction can be computed. The acid of the 
first fraction is then determined and then that of the second calculated and added to it. 

Physiology of the Gastric Juice. — After the ingestion of a test meal the 
secretion of gastric juice begins almost immediately. The hydrochloric 
acid will at first be bound as soon as secreted but by the end of a half -hour 
after the test breakfast or 2 hours after the Riegel meal enough will have 
been secreted that some will remain free. The amount of acid rises to a 
maximum where it remains until the products of digestion pass on into the 
intestine. Then the acidity will begin to fall. The gastric juice as it 
emerges from the gastric glands would seem to contain about 0.5% HC1. 
This is at once partly neutralized by the other constituents of the juice so 
that 1 hour after the Ewald breakfast the total acidity normally averages 
from 40 to 60 A. P. of 0.15% to 0.22% HC1. Lactic acid is not present 
and phosphates are not present in any important amount. Of this total 
acidity the free HC1 will vary from 20 to 60 A. P. (from 0.05% to 0.2%) and 
the bound from 0.012% to 0.11%. With the Riegel meal the normal total 
acidity is about 75 A. P. (from 90 to 100 A. P.) and the free about 44 A. P. 

An acidity per cent, over 70 (0.25% HC1) has been considered hyper- 
acidity. This is far from the truth. The acidity of the normal stomach is 
sometimes 0.33% (A. P. 90) or over and the person quite free from gastric 
discomfort of any kind. We were able to determine this from the analysis 
by our medical students of their own gastric contents which at one time we 
required them to make. " Hyperacidity " is not a question of the percentage 
of HC1 in the gastric juice, but of the contraction of the pylorus which leads 
to slight retroperistalsis and this causes the symptoms interpreted as 
" hyperacidity " (pyrosis, acid eructations, etc.) which may be present 
when the acidity per cent, is normal or slightly low. 

The Value of the Tests for Acidity of the Stomach. — Of all the above 
tests that for the presence of free HO is the most important. A gastric 
juice which contains a normal amount of free HC1 will quite certainly be 
normal in other ways, since the secretion of this acid is the first gastric 
function to suffer in any disturbance of this mucosa. While practically 
every stomach can secrete some HC1 the normal organ will always provide 
an excess, that is, some free HC1. The quantitative determination of the 
total acidity and of the free HC1 is of greatest value not in diagnosis but 
in following the progress of our therapy. Very accurate work is clinically 
of little value since the total amount of secretion can never be recovered. 



350 CLINICAL DIAGNOSIS 

If a bread and water meal is used the phosphates may be disregarded. 
If free HC1 is present organic acids may be disregarded (provided the stom- 
ach was clean before the meal and none present in the food). If the total 
acidity is high and no free HC1 present, the acidity is due for the most part 
to organic acids. This may be confirmed by the lactic acid test or the odor 
of the other organic acids. Many bacteria will be present. If free hydro- 
chloric is present and the total acidity low, the acidity will be due to hydro- 
chloric acid and the motility of the stomach may be assumed to be good 
since the acid-binding bodies have passed on into the duodenum. If, on 
the other hand, the total acidity be moderate and free acid small in 
amount a poor motility may be assumed with the retention of the prod- 
ucts of digestion. 

A review of past work using Ewald's breakfast and i specimen of gastric contents 
is given by Sahli as follows: (A) There is normal acid secretion: (i) Often in ulcer of 
the stomach and stenosis due to the contraction of its scar; (2) in gastric neuroses; and 
(3) in simple atony. 

(B) Hydrochloric acid is over 0.2% (A. P. 55) and the total more than 70 A. P. (it 
may reach 0.35% and very rarely 0.8%) : (1) In the majority of cases of ulcer of the stom- 
ach; (2) in true continuous hypersecretion (but not the hypersecretion ■ due to motor 
stasis) ; (3) in simple hyperacidity and hypersecretion during digestion, at which time the 
per cent, of acid may be abnormally high; (4) in paroxysmal hypersecretion (gastroxynsis) 
in neurotic individuals, who, following some excitement or other disturbance, vomit 
large amounts of acid juice; (5) in some cases of chlorosis (in 22 of 30 of Riegel's cases); 
(6) in early stages of chronic gastric catarrh; and (7) often in insanity. 

(C) The secretion of hydrochloric acid is diminished in (1) fevers; (2) in severe 
anemias; (3) in the majority of cases of chronic gastric catarrh; (4) in many gastric 
disorders due to general neuroses; (5) in many forms of mental diseases; (6) after long- 
standing jaundice; (7) in many chronic cachexias, as tuberculosis of the lung, but not 
always; (8) in chronic passive congestion due to heart disease or to emphysema, etc.; 
(9) sometimes in chronic nephritis; (10) after the long use of alkaline and saline purges; 
(n) as a " fatigue " symptom following periods of hypersecretion. 4 

(D) Free hydrochloric acid is absent on repeated examinations (and yet the stomach 
always contains a certain amount of this acid bound) in all conditions under C of a 
severe grade (especially in amyloid disease of the stomach, toxic gastritis, nervous 
dyspepsia, phthisis, and cardiac disease). Its absence is most important in (1) severe 
febrile diseases, particularly the infections; (2) gastric carcinoma (also other carcino- 
mata) ; (3) atrophic gastric catarrh; (4) pernicious anemia. The most important of these 
is gastric carcinoma. 

Standards vary with nationalities, with classes of society and still more with indi- 
viduals. Strauss, at Giessen, thought 68 a fair average total acidity; at Berlin 47. In 
this country we use an unusual meal and must judge the patient not according to any 
physiological normal, but to an empirical standard gained from the examination of 
many cases. 

Among 526 cases of the Johns Hopkins Hospital clinic whose records were studied 
were the following. In all cases the Ewald breakfast was used. 

Pernicious Anemia, 13 Cases. — Amount removed, 10 to 80 c.c. All were subacid, 
the highest total acidity being 38 (A. P.) and below 10 A. P. in 10 cases. In only 1 was 
there any free HC1. In 2 the fluid was neutral to litmus; in 1, alkaline. Lactic acid was 

4 Foster and Lambert, Jour, of Exp. Med., 1908, vol. x, No. 6, p. 820. 



THE VOMITUS AND GASTRIC CONTENTS 351 

present in 2 cases (in 1 of these the diagnosis was confirmed at autopsy, in the other, not). 
In 2 cases of severe secondary anemia the fluid was only slightly subacid and free HC1 
was present in both. 

Malignant Disease not of the Stomach. — Of these cases, 10 were carcinomata, and 4 
sarcomata. All were subacid (total acidity less than 40) . Of the carcinoma cases free HC1 
was present in 7, was absent in 2 and the fluid neutral to litmus in 1. Of the sarcoma 
cases in none of the 4 was any free HC1 present. 

Catarrhal Jaundice, 9 Cases. — The fluid removed varied in amount from 10 to 86 
c.c; total acidity from 10 to 70 A. P. In 3 cases no free HC1 was present; in 1 the fluid 
was alkaline to litmus; in none was lactic acid found. In a few cases the acidity pro- 
gressively diminished during the course of the disease. 

Cholelithiasis, 14 Cases. — Amount removed, 5 to 120 c.c. There was hyperacidity 
in 1 case (total 79 and 82 on 2 examinations, free HC1, 42 and 49 respectively); 
normal acidity (40 to 70) in 6 cases ; below 40 in 6. In 4 of these 6 there was no free HC1. 
Lactic acid was demonstrated in 1. 

Cirrhosis of Liver, 6 Cases. — Normal acidity was present in 1, subacidity in 5. In 4 
of the 6 cases no free hydrochloric acid was present. Of these 6 in 1 the gastric juice 
was practically neutral, while that of another of these cases contained lactic acid. 

Tuberculosis of Lungs, 10 Cases. — The total acidity was normal in 3 of these 10 
cases and subacid in 7. In 3 there was no free acid. Of these 3 the fluid was neutral in 
1 and 1 contained lactic acid. 

Intestinal Troubles. — Diarrhea, q Cases. — In 4 the acidity was normal, in 4 subacid 
and in 1 almost neutral. In 4 no free acid was present; 1 contained lactic acid. Con- 
stipation, 5 cases, of which 2 had normal acidity, 3 were subacid, and in 1 there was no 
free hydrochloric acid. Colitis, 2 cases, both subacid and without free acid, and both 
containing lactic acid. Amebic dysentery, 1, which was hyperacid (92 total and 83 free). 

Arteriosclerosis and Cardiac Diseases, 17 Cases. — Of these 8 were normal, 6 subnormal 
and 3 without free acid. One of these was almost neutral. 

In a group of 36 cases of miscellaneous diseases, 25 showed normal gastric conditions. 
Subacidity without any free acid was present in cases of heat prostration, of enteroptosis, 
chronic bronchitis, peripheral neuritis (with lactic acid), chronic nephritis, broncho- 
pneumonia, and malaria. 

Fractional Determination of Gastric Secretion. — The inaccuracies 
inherent in the usual method of examining the gastric contents removed 
just once and therefore seldom at the height of digestion, have led to the 
development, by Rehfuss 5 of a technic using a modification of Einhorn's 
duodenal tube which makes it possible to examine several specimens 
removed at intervals during the digestion of a single meal. The test meal 
consists of 40 gms. of water crackers and 10 oz. of water, 2 of which are. 
reserved to be swallowed with the tube. The patient is allowed about 
10 minutes to consume this meal and then immediately swallows the tube 
together with the 2 oz. of water. The patient should be careful not to 
swallow the saliva which forms rapidly during the next few minutes. The 
patient is made as comfortable as possible, is urged to read, etc., and suffers 
little inconvenience with the tube in his mouth. At 20-minute intervals 
from 15 to 25 c.c. of the stomach contents are aspirated by gentle suction 

5 Am. Jour. Med. Sci., June, 1914, vol. 147, p. 848; Jour. A. M. A., Sept. 12, 1914, 
p. 909. 



352 



CLINICAL DIAGNOSIS 



until no more can be obtained when the patient is in the dorsal, right and 
left lateral and knee-chest postures. Each fraction is examined separately 
as regards free and total acidity and the presence of pepsin. 

Fishbaugh 6 using Rehfuss' method calls attention to the curves of 
gastric secretion which the cases studied by this method present. In i 
group of cases the curves of the secretions reach their maximum in 91 
minutes (average) and fall towards the end of gastric digestion. This 




Chart I. — M. H., Duodenal ulcer. Ewald breakfast given November 22, 1920, without previous 
lavage and removed through a Rehfuss tube at 15 minute intervals. That evening the stomach was well 
washed out and the breakfast repeated. See Chart II. The tube had not entered the duodenum in 

3 hours. 

would seem at first thought to be the normal condition. The need of 
further secretion past the stimulus lessens, the juice is less concentrated, 
while some of the acid is neutralized by the products of digestion. In a 
second group the curves are still rising at the end of gastric digestion (aver- 
age 127 minutes) so that the last few cubic centimeters of gastric juice are 
the most concentrated of all. This is normal in some healthy individuals. 
In the third group of cases with gastric secreti on absent or delayed there 
6 The Jour, of A. M. A., Oct. 28, 1916, vol. lxvii, p. 1275. 



THE VOMITUS AND GASTRIC CONTENTS 



353 



may be an absence of acid and enzymes, or an absence of acid with enzymes 
present, or the acid and enzymes may appear late. 

To illustrate this method and also to emphasize some of the mistakes inherent in 
clinical gastric analysis we give 2 charts of examinations made on 1 patient on successive 
days. The patient, a man 41 years old, is a clear case (the clinical history and especially 
the rontgenological examination positive for this) of duodenal ulcer with considerable 
retention. A test was made in the afternoon of Nov. 22, 1920. Chart I. The meal 
was given at 12.30 p.m. and 8 specimens removed at 15-minute intervals beginning 




Chart II. — M. H., November 23, 1920. Ewald breakfast given at 7-30 a.m. and removed at 15-minute 
intervals. The stomach had been well washed the evening before. Compare with Chart I. Between 
9.30 and 9.45 the tube entered the duodenum. 

at 1 .00 p. m. The total acidity was high reaching 124, the free acidity reached 74 while 
the bound acidity was very high, even from 50 to 64. This shows that there was still 
considerable food in the stomach before the meal was given. 

That night the stomach was well washed out and the same meal given at 7.30 a.m. 
the following morning. Chart II. In 1 hour the free acidity was 56 and the total 72. 
According to former methods therefore this would pass as a case of normal acidity. 
Thirty minutes later however the free was 94 and the total 104. 

The former methods of examination (removal of the meal in 60 minutes) 
led to frequent error in the cases of the first 2 groups, but in the third to 
23 



354 CLINICAL DIAGNOSIS 

very frequent and serious errors in that these cases were usually diagnosed 
" achylia gastrica " which is seldom the true condition. It is possible that 
in those cases with late secretion the mucosa is fairly normal but responds 
only to the chemical stimulus, the psychic being absent or ineffectual. 

Undoubtedly the work of Rehfuss and his co-workers is one of the great 
advances in clinical medicine. The results of gastric chemistry had been 
so unsatisfactory that the diagnosis of our gastric cases has been left almost 
entirely to the rontgenological departments, although in the very nature 
of the case their aid, while valuable, is of necessity limited and most of the 
evidence they obtain indirect. 

Sahli's Desmoid Reaction 7 is based on the assumption that catgut in 
its raw state is soluble in gastric juice, but indigestible in pancreatic juice. 

A pill of methylene blue, 0.05 gm., or of iodoform, 0.1 gm., or of both, and sufficient 
ext. glycyrrhizae to make a mass not over 3 or 4 mm. in diameter is enclosed not too 
tightly in a square piece of thin rubber dam the twisted neck of which is tied with 3 turns 
of raw catgut, number 00, previously soaked until soft in cold water. Both knots should 
be on the same side of the bag. The free edge of the rubber should extend about 3 mm. 
beyond the ligature, and its edges should not cohere. The completed pill should sink 
instantly in water and should be proven to be water-tight. 

This " desmoid pill " is given with or just after the midday meal and the urine, 
collected at intervals of 5, 7, and 18-20 hours, examined for the methylene blue (or 
iodine, or both). To demonstrate the methylene blue it may be necessary to boil the 
urine with % volume of glacial acetic acid. The iodine is recognized by the rose color 
which develops when pure nitric or sulphuric acid is added and the urine then shaken 
out with a little chloroform. If these indicators are found within frcm 18 to 20 hours 
after the pill is swallowed the test is positive; otherwise, negative. 

It is claimed that this test is simple, comfortable for the patient, and that it will 
test the digestive (both HC1 and pepsin) activity of the stomach at the height of digestion 
of a full meal and therefore sometimes indicates good functional activity when the test 
breakfast shows no free HC1. It is useful therefore to distinguish between true achylia, 
as in carcinoma or pernicious anemia, and that present with less serious disorders in 
which the stimulus of the Ewald breakfast is insufficient. 

The most important result obtained thus far is that in no case of cancer of the 
stomach or of pernicious anemia in which the test breakfast showed an absence of free 
HC1 did the capsule open. This test has a real though limited value, but it cannot 
entirely replace the stomach-tube and test meal, or the X-ray. 

Various other methods of testing the gastric juice without using a tube have been 
proposed, especially by means of a hollow capsule containing reagents. The best of 
these was described by Rehfuss. 8 

The ferments of the gastric juice are pepsin, rennin and lipase. Riegel 
believed that much of the attention which has been directed to gastric 
acidity should be turned to the study of gastric ferments. 

Pepsin. — The qualitative determination of pepsin need never be made 
when there is any free hydrochloric acid present, since the pepsin-forming 
function of the stomach is more resistant in disease than is the hydrochloric- 

7 Sahli, Corresp. Bl. f. Schweiz. Artzte, 1905; Boggs, Johns Hopkins Hosp. Bull., 
1906, vol. xvii, p. 313; Carey, Boston Med. and Surg. Jour., May 2, 1907, vol. clvi, 

P- 563. 

8 Am. J. Med. Sci., June, 1914. 



THE VOMITUS AND GASTRIC CONTENTS 355 

acid-forming function. (The water-secreting function is the most resistant 
of all.) The determination of pepsin is valuable when there, is no free HC1, 
as in carcinoma, pernicious anemia and atrophic gastric catarrh. Schiff 
found that in cases with marked hypacidity and anacidity due to benign 
causes the amount of pepsin secreted is little changed, but that in slight 
hypacidity due to gastric cancer it may be greatly reduced. 

Qualitative Determination. — The presence of pepsin is assumed if the 
gastric juice (HC1 added if none be free) will digest egg albumin or fibrin. 
The fibrin is prepared as follows : Fresh ox-blood is whipped and the fibrin 
kept in running water until perfectly colorless. It is then cut in fragments 
of equal size which are put for a few days in alcohol and then for i or 2 
days in a cool, concentrated, neutral, carmine solution until fully stained. 
It is then well washed, pressed out and kept in glycerin stained with car- 
mine. Before use the fibrin is well washed with water to remove all the 
glycerin . To prepare egg albumin the egg is boiled for 5 minutes, not longer, 
and the white then cut with an ordinary cork-borer into 5 mm. cylinders 
and these into disks 1 mm. thick. These disks are kept in glycerin and are 
well washed in water before they are used. 

To the fluid to be tested for pepsin hydrochloric acid is added if neces- 
sary until some is free, then the fibrin or egg disks are added and the speci- 
men put in a thermostat. If pepsin is present the fibrin will show signs of 
digestion first by the liberation of the carmine in from 15 to 30 minutes, 
the egg albumin first by the rounding of the disks' edges in from }{ to 4 hours. 

Sahli recommends that both albumin and fibrin be tried, since some 
gastric juices can digest the fibrin easily but not the albumin, and that the 
tubes be kept at room temperature in order that the difference in them 
may be more clearly observed. 

Quantitative Determination. — The general law of ferment action is that 
activity varies approximately as the square root of the amount of the fer- 
ment present. 

Mett's quantitative method of estimating the amount of pepsin is 
fairly accurate if the gastric fluid has been diluted 16 times. The fresh egg 
albumin is filtered and exposed to a vacuum produced by a suction pump 
for several hours in order to remove all gas. A beaker is then filled with the 
albumin, a bundle of glass tubes, each about 1 cm. long and 1 to 2 mm. 
wide, are then immersed in it and the beaker heated for 5 minutes in water 
at 95 C. The tubes are then carefully removed, their outside cleaned, 
and both ends closed with sealing-wax. One cubic centimeter of the filtered 
gastric juice is mixed with 15 c.c. of 0.05N HC1 and well shaken. Pieces 
of the tubes full of albumin 2 cm. long remain in this fluid for 24 hours in 
the thermostat. The average distance to which digestion has progressed 
into the tubes is then measured. The square of this multiplied by 16 will 
give the units of pepsin in the gastric juice. (By unit is meant the amount 
of pure pepsin which in 24 hours will digest an average of 1 mm. of the albu- 



356 CLINICAL DIAGNOSIS 

min in several tubes.) The distance to which theoretically pure pepsin 
would progress into the tube is 4 mm. 

The Fat-splitting Ferment — Volhardt's Method. — The yolk of 1 egg is mixed 
with 30 to 40 c.c. of water; 10 c.c. of this mixture are added to the gastric juice, both 
fluids having been warmed separately in the thermostat. The mixture is then kept for 
2 hours in a thermostat which registers from 37 C. to 40 C, then cooled. Ether, 75 c.c, 
and a few cubic centimeters of alcohol are then added and the specimen Yvrell shaken. 
A measured amount of the fat-containing ether extract thus obtained is now mixed with 
50 c.c. of neutral alcohol and titrated with 0.1N NaOH using phenolphthalein as indi- 
cator, to determine the amount of fatty acid split from neutral fat by the gastric lipase. 
One next adds 10 c.c. of 0.1 N NaOH and places the specimen on a water-bath for 2 hours 
in a flask connected with a condenser and a calcium oxide tube to exclude C0 2 (or it is 
allowed to stand for 24 hours in a closed flask at room temperature) to saponify the 
unsplit fat. Ten cubic centimeters of 0.1N HC1 are now added to free the fatty acid and 
the mixture is again titrated with 0.1N NaOH, phenolphthalein as indicator, to deter- 
mine the fat which was unchanged by the ferment while the specimen was in the thermo- 
stat. From the relation of these values the percentage of neutral fat which was split 
by the ferment is reckoned, and hence the number of units of ferments present, using. 
Stade's formula, p = -y/f X VM n which p equals the product of digestion,/ the units of 
ferment, and t the time. If "/ " represents the amount of ferment which will split 1 % 
of the fat in 1 hour, then, if after 3 hours one finds 6% split, 12 units of ferment must 
have been present. 

Volhardt found that in the stomach in 2 hours from 30 to 36% of the neutral fat 
is split. The fatty acids thus liberated and later dissolved in the bile evidently aid in 
the emulsion of the neutral fats of the foods. It is possible therefore that the stomach 
does considerable of the work usually attributed to the pancreas. 

It has been found that in hypochylia and achylia the lipase either is diminished in 
amount or is absent. 

Rennin. — The presence of rennin in the gastric juice is proved if this secretion, 
after it has been neutralized, coagulates milk without change of reaction. 

RiegeVs method of determining the presence of rennin is to mix from 5 to 10 c.c. of 
gastric juice (neutralized with 0.1N NaOH) with from 5 to 10 c.c. of fresh milk and to 
place this mixture in a thermostat. If rennin is present coagulation will be evident in 
from 10 to 15 minutes. If a longer time is required one should exclude curdling by some 
lactic acid which has been formed. 

A quantitative determination of rennin may be made by mixing, in a series of tubes, 
equal volumes of fresh milk and various dilutions of the neutralized gastric juice and 
noting the highest dilution of the juice which will coagulate the milk. Boss found in 
normal cases this to be a dilution of 1 : 100 to 150. In cases of hypochylia the ability 
of the rennin to coagulate the milk may disappear when the juice is diluted 1 : 5 to 10. 
Glassner found that in some cases of pyloric cancer the rennin secretion is normal while 
that of pepsin is diminished, while in cancers of the fundus both rennin and pepsin are 
diminished. In general the variations in the amount of rennin run parallel to those of 
pepsin. Since rennin is the easier of the 2 ferments to estimate it may be that in time 
we shall follow its secretion rather than that of pepsin. 

The Products of Gastric Protein Digestion. — We may divide the 
products of the peptic digestion of proteins into the following groups: 
albumoses, peptones and the products of further cleavage. The soluble 
albumin is precipitated by heat, the albumoses by the addition of an equal 
volume of saturated zinc sulphate and the peptone by phosphotungstic 



THE VOMITUS AND GASTRIC CONTENTS 357 

acid. The filtrate after this last precipitation will contain all products 
below the so-called peptone stage. Those various fractions of digestion- 
products are best determined by estimating the nitrogen in each filtrate, 
care being taken to reduce all quantities into terms of the original volume 
of the stomach contents obtained. 

Benedict 9 advises to determine these precipitates volumetrically after 
centrifugalizing them to their smallest possible volumes. The meal used 
should contain but i proteid and this should be carefully weighed. We 
have used nutrose (a casein preparation) with good success. The nitrogen 
fractions of the contents of the fasting stomach of each patient studied 
should also be determined and these corrections made. The meals should 
be given at the same hour of the various days, and removed at the end of the 
same period of time. 

From a theoretical point of view it is interesting that in cases of carcinoma of the 
stomach the digestion of proteid is so much more rapid than normal that one must 
assume the presence of an abnormal ferment. 10 Artificial digestion experiments using 
as ferment heated and unheated carcinoma tissue strengthen this assumption. 

In our benign cases studied in this way the Ewald breakfast being used the average 
amount of nitrogen in albumose found was 51.7%; in the phosphotungstic acid precipi- 
tate, 31.4%; in the residue, 16.9%. In the carcinoma cases these figures were respec- 
tively 27.5, 47, and 27.6%. 

Starch Digestion. — Undoubtedly the current idea that the saliva is not 
important in. starch digestion and that the pancreas does it all must be 
corrected since from 50 to 70% of the starch of the food is rendered soluble 
in the stomach. The ptyalin digestion can continue throughout the stom- 
ach contents until the total HC1 reaches 0.12% and within the food masses 
for a much longer time until the acid has penetrated to their center. Hence 
starch digestion is reduced in cases of hypersecretion and hyperacidity. 

The stages of starch digestion are: soluble starch, erythrodextrin, achroodextrin and 
maltose. The relative amount of these may approximately be detected by the use of 
a very weak Lugol solution. The colors obtained vary from the blue to the blue- violet 
of the starches, the red to the mahogany-brown of the erythrodextrin while the iodine 
combination with achroodextrin is colorless. Since the later products of starch digestion 
have a greater affinity for the iodine than the earlier, 1 drop of weak Lugol's solution 
added to a small amount of gastric juice will give no blue color with the starch present 
if much achroodextrin also is there. On the other hand, if very little of the higher prod- 
ucts are present the starch will give its blue color. From the number of drops of the 
Lugol solution which must be added before the blue color appears one may estimate 
approximately the extent of the starch digestion. (For a more accurate method see 
page 383.) 

Lactic Acid. — The presence of lactic acid in the gastric contents is 
significant only when the food eaten contained none and no constituent 
from which it might easily be formed and provided the stomach had been 
well washed out the evening before. Riegel says its presence in the stomach 

9 Am. Jour. Med. Sci., 1904, vol. cxxvii. 
10 Emerson, Deutsch. Arch. f. klin. Med., vol. lxxii, p. 415. 



358 



CLINICAL DIAGNOSIS 



in never normal except during sugar digestion. There are bacilli in the 
mouth which can produce lactic acid but not in the time allowed by a 
test meal. For this reason the safest test meal so far as lactic acid is con- 
cerned is Dock's which consists of water and a shredded wheat biscuit. 
Uffelmann's test for lactic acid is the one in com- 
mon use. To about 20 c.c. of 1% carbolic acid made up 
fresh each time in a test-tube is added i-drop of 10% 
Fe 2 Cl 6 . A deep amethyst color is produced. Distilled 
water is now added until the blue color is so faint that 
one can see through the tube. This fluid is then divided 
in 2 test-tubes of equal diameter. To the 1 is added a 
drop or 2 of the gastric juice to be tested, to the other 
the same amount of distilled water. If lactic acid be 
present the amethyst color will change to a definite 
yellowish-green (canary-green) color. 

The decolorization of the amethyst-colored fluid is 
not the test, but the development of a definite yellow. 
The blue is merely for contrast and so one may dispense 
with the carbolic acid. It is well to control the test 
with dilute lactic acid. 

The test proposed by Strauss is better. In a small, 
specially marked separating funnel (see Fig. 70) one 
mixes 5 c,c. of the gastric juice and 20 c.c. of alcohol- 
free ether. One drop of hydrochloric acid is added to 
liberate any lactic acid which is bound to proteid and 
( : S CT ] the tube well shaken to extract the lactic acid. The 

gastric juice is then allowed to run out and 5 c.c. of 
distilled water added to the ether extract, then 2 drops 
(from a medicine dropper to insure uniform size) of a 
1% Fe 2 Cle solution. If at least 0.1% of lactic acid is 
present the water layer will take a definite canary-green 
color. This test is perhaps not as delicate as the other, 
but if it is positive it may be said confidently that a 
pathological amount of lactic acid is present. It is 
raUng 7 °f^nei a fo? factic desirable to extract the gastric fluid with ether since a 
suggestive color may be given by sugar, proteid and 
alcohol and by some other inorganic acids (oxalic, citric, tartaric, etc.) 
while a positive test for lactic acid may be prevented if much phosphates 
or peptones are present. Again, ferric chloride may so cloud the gastric 
juice that the test will be obscured. 

This test has been further modified by Knapp u who extracts 1 c.c. of gastric juice 
with 5 c.c. of ether. He then superimposes this extract upon a freshly prepared 1 : 2000 
ferric chloride solution. Lactic acid will be indicated by a canary-colored ring. 




New York Med. Jour., August, 1901. 



THE VOMITUS AND GASTRIC CONTENTS 359 

De Jong 12 adds to 5 c.c. of the gastric juice 1 or 2 drops of HC1 and evaporates this 
to a syrup over a free flame. He then extracts the residue with a little ether. The volume 
of extract is then made up to 5 c.c. with distilled water, 1 drop of 5% Fe 2 Cl 6 added, and 
the whole well shaken. A definite green color is produced by 0.05% lactic acid. 

Quantitative Determination of Lactic Acid. — For clinical purposes the Strauss modi- 
fication of the Uffelmann method is an accurate enough estimation of the amount of 
lactic acid present since if that is positive the amount certainly is pathological and this 
is all we need to know. 

Other Organic Acids. — Acetic, butyric, and valeric acids may be met with in the 
gastric contents and be recognized by their odor. They are the result of the bacterial 
decomposition of foods. Why in some cases one, in others, another, acid is formed is not 
known. One of our patients who gave all the symptoms of hyperacidity recently was 
given an Ewald test meal and at once complained of the taste of vinegar. The gastric 
contents were found rich in acetic acid. 

Acetic acid carefully neutralized with soda if tested with 1 or 2 drops of Fe 2 Cl 6 
solution will give the bluish-red color of ferric acetate. 

Butyric acid in the presence of fine fragments of CaCU separates in fine oily droplets. 

Total Organic Acid — Hehner-Maly Method. — The principle of the Hehner- 
Maly method for determining the total amount of organic acid in the gastric contents is 
that if a mixture of organic and inorganic acids is ashed the inorganic salts will remain 
neutral, while the organic will be changed to alkaline-reacting carbonates. The original 
acidity minus the final alkalinity may be considered to equal the mineral acidity. 

Ten cubic centimeters of filtered gastric juice are neutralized with 0.1N NaOH, 
using phenolphthalein as an indicator (amount necessary = a). This fluid is then evap- 
orated and ashed, the ash taken up with water and titrated with 0.1N HC1, phenol- 
phthalein again used as indicator (amount necessary = &) , a — b =the mineral acidity ( =c) 
then a—c = organic acidity. 

Bases of the Gastric Juice. — Sodium is the most important base of the gastric juice. 
Mehring has shown that the stomach secretes sodium carbonate to control the amount 
of hydrochloric acid. Reissner showed an increased secretion of alkali in gastric cancer. 
Ammonia also is present in considerable amount (normally 0.1 to 0.15 p.m.). Of all the 
tissues of the body which have been examined, the gastric wall contains the most ammo- 
nia. This throws considerable doubt on the accuracy of all titrations of gastric contents 
in which phenolphthalein was the indicator used (see page 346). 

Fermentation. — Two kinds of fermentation may take place in the 
stomach, 1 with gas formation and 1 with lactic acid but no gas formation. 
Neither occurs unless considerable stasis is present. The former is the 
rule if free HC1 is present, the latter when this free acid is much diminished 
in amount or is absent. 

Lactic acid fermentation is proved by the presence of this acid in the 
gastric contents provided foods can be excluded as the source of the acid. 
Small amounts may be found when there is no genuine stasis as "after a 
heavy meal or if there are pockets or little clefts in the stomach wall. 

To test for gas fermentation 2 fermentation tubes should be filled with the well 
shaken unfiltered gastric contents. To 1 of these tubes, which serves as a control, is 
added a little glucose (since all the carbohydrate of the material tested may already 
have been fermented). The tubes are then left in the thermostat for 24 hours (some say 
for 3 or 4 days) at the end of which time the presence of gas, if any, is noted. Both 

12 Arch. f. Verdauungskrankheiten, Bd. 2. 



360 CLINICAL DIAGNOSIS 

carbohydrates and proteids may ferment in the stomach. This test is of some value 
in determining the degree of gastric stagnation; that is, the abundance of the gastric 
flora. The organisms of fermentation which are found in the stomach are yeasts, 
sarcinse and long bacilli. Sarcinas may be present in. cases of high acidity, bacilli in 
cases of low acidity and of anacidity. Some believe that the presence of sarcinae in 
cases of low acidity, or of long bacilli in cases of reduced acidity indicate cancer of the 
stomach. Yeasts are common in gastric contents. Coyon doubts that sarcinas explain 
much of the fermentation, but he has isolated 2 bacilli, the enterococcus and Coccus 
radiaire, which he thinks are more important. 

Among the products of fermentation have been found formic, lactic, and acetic 
acids, the higher fatty acids, ethyl alcohol, aldehyde, ammonia, hydrogen disulphide, 
and carbon dioxide. 

Hydrogen disulphide, from the decomposition of proteid, is rarely found in the 
stomachs of patients with malignant, but frequently in the stomachs of patients with 
benign, stenosis of the pylorus. The saliva and some foods, as radishes and onions, must 
be excluded as its source. This gas may be recognized by its odor, or by suspending 
over the fluid a strip of paper moistened with alkaline sugar of lead. A positive test 
does not suggest the presence in the stomach of any particular organism. 

The gastric sediment should be studied with care since it often yields 
evidence of value in diagnosis. 

Lyon's 13 method is as follows. He uses a small metal capsule 1.5 cm. 
long by 6 mm. in diameter with a shaft 4 mm. long by 3 mm. in diameter, 
to which a small capillary rubber tube is securely fastened by a silk thread. 
The capsule is perforated at the extreme tip with a hole 1.5 mm. in diameter, 
in a line continuous with the caliber of the tube, and the body of the capsule 
is similarly perforated with 8 additional holes 1.5 mm. in diameter. To 
facilitate cleaning the capsule is made in 2 parts which unite by a screw 
thread. The capillary rubber tube is 1 meter long and of various calibra- 
tions, but should be at least 3 mm. in diameter. This tube can be readily 
swallowed with minimal discomfort and can be left in situ for several hours 
to admit of fractional analysis of the gastric juice after an Ewald breakfast, 
or allowed to pass into the duodenum for the recovery and analysis of 
duodenal or jejunal contents. 

The patient swallows the capsule and tube on an empty stomach, prefer- 
able in the early morning fasting state. Then by means of a 1- or 2 -ounce 
aspirating syringe, with a capillary tip and a close-fitting asbestos plunger, 
gentle aspiration is made to recover the residual contents of the fasting 
stomach. From 100 to 150 c.c. of plain, warm water is next introduced 
by means of the syringe and is gently aspirated and then forced back again 
into the stomach, perhaps a dozen times. The lavage water, which was at 
first macroscopically clear, soon becomes gradually turbid and contains 
variously-sized flocculent bodies ranging from pin-point to 3 to 5 mm. in 
size. After thus douching the -gastric mucosa all of the fluid is aspirated 
from the stomach. A small portion is tested for occult blood and the 
remainder mixed with equal volume of a 10% solution of formalin. This 

13 Am. Jour, of the Med. Sciences, Sept., 1915, No. 3, vol. cl, p. 402. 



THE VOMITUS AND GASTRIC CONTENTS 361 

is then filtered, the filter paper is punctured and the residue washed into 
a clean bottle with the 10% formalin solution and allowed to stand for at 
least 3 hours. The residual contents obtained without lavage is treated in 
the same way. 

The sediment is filtered through a smooth filter paper and the sediment 
washed down to the tip of the filter by means of a wash bottle. The tip 
of the filter paper containing the sediment is then cut off and the paper 
folded so that it encloses the sediment. This package is now wrapped in 
i layer of gauze and tied fast with a thread. This is placed in Acetone I 
for i hour, then in Acetone II for i hour and in Acetone III for 2 hours. 
It is then transferred to paraffin and chloroform, 1 hour, to paraffin 
(M.P., 52 C), 1 hour, paraffin (M.P., 52 C), 2 hours, the paper then 
removed and the mass of sediment imbedded in paraffin and serial sections 
cut and fastened to slides. These are heated in a dry heat sterilizer at 
70 to 8o° C. for 30 minutes or until the sections are perfectly dry. The 
slides are then run through xylol (in Coplin's jars) for 5 to 10 minutes, 
absolute alcohol for 5 minutes, 95% alcohol for 5 minutes, 80% alcohol 
for 5 minutes, and water for 5 minutes. They are then stained in hema- 
toxylin for 5 minutes, washed with water and then 1% eosin and again 
washed. They are next run through 80%, then 95%, then absolute alcohol 
for a few seconds, cleaned in xylol and mounted in balsam. 

In sections from the aspirated residue of the normal fasting stomach one 
finds occasional epithelial cells and occasional leukocytes with protoplasm 
intact in those cases in which chemical titration shows faintly acid or neu- 
tral or slightly alkaline reaction. Boas and Paul Cohnheim have pointed 
out that digested protoplasm of epithelium cells or leukocytes indicates 
the presence of free hydrochloric acid and pepsin. When the protoplasm 
of the epithelial cells is still intact it is possible to differentiate endogenous 
gastric cells from those originating from the mouth, pharynx, respiratory 
tract and esophagus. One frequently encounters the snail-like bodies first 
described by Jaworski which Boas and Paul Cohnheim believe to be mucus 
acted upon by hydrochloric acid. If there has been regurgitation from the 
duodenum there may be crystals of some of the bile salts. 

Pathologically, in the fasting morning stomach one may find remnants 
of food eaten the night before, such as muscle fibers still striated or partially 
digested; starch granules; vegetable-cells; seeds from berries, any of which 
from a 12 -hour fasting stomach is indicative of motor insufficiency due 
either to pyloric obstruction or rarely to advanced atony, although it 
should be remembered that small amounts of food remnants are not neces- 
sarily pathological (crypts of mucosa and cavities in teeth) . Associated 
with this, if one finds sarcinse in numbers or many yeast cells in process of 
germination it would suggest gastric dilatation with stagnation and fer- 
mentation. Sarcince are rarely found in the ectasis of cancer, except in 
those cases of the ulcus carcinomatosus type. 



362 CLINICAL DIAGNOSIS 

Infusoria, Trichomonas hominis and Megastoma entericum may be 
found. These require for their development an absence of hydrochloric acid, 
an alkaline medium and a mucosa with crypts. Mucus from the respiratory 
tract will float, owing to its air content. Microscopically, it is character- 
ized by the alveolar cells and myelin drops which it contains, while columnar 
epithelium indicates its derivation from the gastric mucous membrane. 
Also, in gastric dilatation one occasionally encounters spores and mycelial 
cells from vegetable moulds. Leukocytes, indicative of an inflammatory 
reaction, are present in large numbers in all cases of gastric ulcer, in many 
of the simple forms of gastritis during the inflammatory or congestive stage 
and in cancer of the stomach affecting chiefly the glandularis. Large num- 
bers can be found in some cases of gastric cancer, while in a very few cases of 
gastric cancer and of subphrenic abscess perforating into the stomach the 
stomach may contain even from 60 to 500 c.c. of pus. One would expect 
to find pus in gastritis phlegmonosa and in local abscess of the wall of the 
stomach. Pathologically, a significant finding is the presence of Opper- 
Boas bacilli (see page 378). They are large non-motile bacilli with a some- 
what typical morphological arrangement in long chains and are readily 
differentiated from the Leptothrix buccalis by acting negatively to Gram's 
stain. In gastric sediments prepared as above described they have a 
tendency to arrange themselves in dense masses, interlaced with one another 
and resemble hair-like balls when viewed under a low-power microscope. 

The normal stomach should contain very few bacteria. When the 
bacterial flora is abundant it indicates a distinctly pathological condition. 
The most common normal invaders of the stomach is the Bacillus coli 
group, but the appearance of diphtheroid bacilli, staphylococci and par- 
ticularly various types of streptococci indicates local infection. Here, too, 
one meets with a pathologically increased number of leukocytes. 

It is surprising how often small isolated fragments or flakes of gastric 
mucosa will be recovered by this method. Minute particles, barely of 
macroscopic size, which would readily escape detection in the lavage water, 
may prove to be the one point upon which the correct diagnosis can be made. 
It is often possible from a microscopic study of these bits of mucosa to 
determine from which segment of the stomach they come, whether the 
fundus, prepylorus, or antrum pylori, bearing in mind the anatomical 
distribution of the different types of glands. Microscopically, these minute 
fragments may show only the peripheral portion of the villus extending 
down to various depths through the glandularis, while in the larger frag- 
ments the entire width of the mucosa, at times including the muscularis 
mucosas, will be found. Some are clearly fragments of cancer. Fragments 
of normal gastric mucosa are not infrequently found in the stomach con- 
tents by those who attach a suction pump to the stomach-tube when empty- 
ing this organ. These men claim that the injury to the gastric mucosa 
which such strong negative pressure produces is of no importance, an 



THE VOMITUS AND GASTRIC CONTENTS 363 

opinion which needs confirmation. Others, more gentle with the mucosa 
in their technic, learn much from the bits of mucous membrane whose pres- 
ence indicates that vulnerability present in malignant disease, in chronic 
gastritis and in some cases of Heynoch's purpura with edema of the stomach 
wall (Morris). The fragments of proliferating mucosa thus obtained in 
some cases of achylia gastrica closely resemble bits of carcinomatous tissue. 
Fragments of tumor should be searched for in the morning stomach wash- 
ings of patients suspected to have gastric cancer. In these washings the 
fragments of tumors, the chains of long bacilli and the parasites' eggs are 
the most important finds. 

Microscopic Examination. — The microscopic inspection of the partially 
digested food of a test meal seldom yields any results of value, although 
mucus and pus can best be recognized in this way. Nothing can be learned 
from the inspection of the muscle fibers. 

Infusoria (see page 362) are sometimes found in the gastric contents 

of patients whose gastric juice has for some time ___ 

been neutral or alkaline in reaction. 

In case of gastric stasis with free HC1 present, 
moulds, yeasts and sarcincB will predominate; if 
there is no free HC1, bacteria. Einhorn found in 
the wash-water of certain cases the spores of 
moulds which he thought lodged in crevices of the 
mucosa and might produce the pyrosis and gas- FlG - 71 — Sarcina ventricuii 

45 ^ . x and yeast cells. X 900. 

tralgia present. A few yeasts (Fig. 71) may be 

found in normal stomachs but they are abundant in cases of gastric 
dilatation. Sarcinae (Fig. 71) occur in large numbers in stomachs with 
benign dilatation, occasionally in gastritis, ulcer, and gastric neuroses, but 
rarely in cancer. One observer reported them in such numbers as to 
form plugs obstructing the pylorus. Ehret found many sarcinse in cases 
with intense fermentation of the gastric contents, but no yeasts nor bac- 
teria. He considers that their presence indicates a marked stasis. Two 
sizes may be found, the large and the small. In a recent case with dilated 
stomach in this clinic, sarcince, huge in size, were present in great numbers. 

Blood. — Traces of fresh blood are often present in the gastric contents. 
It may come from the esophagus, nose, mouth or lung, but more commonly 
its presence is explained by the slight lesions of the pharynx, esophagus or 
stomach produced in vomiting or by the tube. Blood in large amounts is 
found in cases of ulcer, some cancers and from rupture of venous dilatations 
at the cardiac orifice, as in portal obstruction. If the stomach was empty 
at the time of the hemorrhage the blood may appear arterial, but as a rule 
it is dark because of the hematin produced by the gastric juice, is clotted 
and is well mixed with food. 

Definite gastric hemorrhages occur also in chronic passive congestion; 
in cirrhosis of the liver; in various constitutional diseases in which the reason 



364 CLINICAL DIAGNOSIS 

for the bleeding is not apparent, as in the anemias, hemophilia and Hodg- 
kin's disease; in active hyperemia of the gastric mucosa, as in vicarious 
menstruation; and following abdominal operations, especially such as 
involve the omentum, in which case they are a disturbing symptom but 
are of no moment. All the blood of even a profuse gastric hemorrhage 
may pass by the stools. 

In carcinoma of the stomach, in which a slight constant oozing from the 
tumor is the rule, the blood is well mixed with the food, is digested and 
looks like coffee grounds. This in a case of gastric dilatation always sug- 
gests cancer and yet may be present also in cases with hyperacidity and 
hemorrhagic erosions of the mucosa. Such blood must be recognized 
chemically. Blood may arise also from tuberculous ulcers; from slight 
injuries to the mucosa; from small aneurisms of the gastric arteries; and 
from infarcted areas. 

Occult hemorrhages, that is, hemorrhages so slight that the presence 
of blood would not be suspected by inspection but must be detected chemi- 
cally, are common in several conditions which Boas and Kochmann 14 
classify as follows: Those in which the blood is never evident, as gastritis 
anacida, subacidity, hypersecretion, benign ectasis; cases with at times 
gross hemorrhages, especially ulcer; cases with evident blood as a rule, 
cancer especially. 

Deen's Test for Occult Blood. — To the gastric juice are added i c.c. of 
fresh tincture of guaiac and i c.c. of Huhnerfeld's solution (glacial acetic 
acid, 2; distilled water, 1; oil of turpentine and alcohol, of each 100 c.c). 
The mixture is well shaken. If blood is present the fluid will turn blue. 
The test is also given if iron compounds and some vegetables are present, 
hence it had chiefly a negative value. Weber's modification is recommended 
by Riegel. To the gastric contents is added % its volume of glacial acetic 
acid and it is shaken out with ether. After the ether extract has cleared, 
to a few cubic centimeters are added 10 drops of guaiac tincture and 20 to 
30 drops of turpentine. Blood will give a bluish- violet color. In this case 
only raw or rare-cooked meat are to be excluded. 

For the spectroscopic test, the gastric contents are diluted with water, 
a few drops of concentrated acetic acid added and it is shaken out with 
% its volume of ether. In a few minutes a clear layer of the brown ether 
solution of hematin will be obtained. Since the 4-band spectrum of hematin 
in acetic acid could be due also to chlorophyll, an alcoholic solution of 
KOH is added and the hematin reduced with (NH 4 ) 2 S. The resulting red 
fluid will give the 2 -line of reduced hematin. 

Motility of the Stomach. — Any disturbance of the motility of the 
stomach is of far more importance than are disturbances of secretion. 
If the motility is good the persori may live for years in ignorance of the 
fact that he has no gastric juice. But if motility be impaired the symptoms 

14 Arch. i. Verdauungsk., 1902, vol. viii, Heft. 1, 2. 



THE VOMITUS AND GASTRIC CONTENTS 365 

will be severe and the stagnation of food in the dilated stomach will soon 
produce serious results. It is of interest that the symptoms of hyperacidity 
(heart-burn, acid eructations, the vomiting of acid fluid, etc.) are due to 
disturbed motility and bear little direct relationship to the actual degree 
of gastric acidity. 

The most rapid motility is seen in cases of jejunal fistula high up. In 
these cases of starvation the stomach seems to hurry the food into the 
intestine at as rapid a rate as possible. 

It is quite important in all diagnostic work to wash the stomach well 
in order to be sure that it is empty, since in cases of achylia the tube may 
siphon nothing and yet the wash- water show considerable solid matter. 

Megalogasiria means enlargement of the stomach. This may or may 
not be accompanied by motor insufficiency. 

Ectasis refers to enlargement with motor insufficiency. The term 
" atony " is used if it is due to real weakness of the muscle wall; " hyper- 
tonic ectasis " when due to pyloric stenosis. 

Motor insufficiency of the stomach may be absolute or relative and 
the resulting dilatation is in general due to i of 2 factors: (1) To atony of 
the gastric wall, in which case the muscle is not strong enough to empty 
the stomach; in this group are found the largest stomachs. Strauss 15 
reported 1 with a capacity of 5K liters. (2) Relative muscular insufficiency. 
By this is meant that while the muscularis may be abnormally strong and 
hypertrophied an obstruction at the pylorus renders the exit of food diffi- 
cult. As long as the hypertrophy of the muscle can compensate for the 
obstruction there will be no dilatation of the stomach. 

A pyloric stenosis may be congenital or acquired. Acquired stenosis 
may be due: to the contraction of scars of ulcers; to cancers; to hyper- 
trophic stenosis (said to be the result of a continued pyloric spasm stimu- 
lated by hyperacid gastric juice or by an ulcer) or to scars resulting from 
irritating poisons. Or, the obstruction may be due to the pressure of neigh- 
boring tumors ; to twists ; to diverticula ; and, finally, to malpositions of the 
pylorus due to adhesions. 

Disturbances of gastric motility explain most of the gastric symptoms 
usually ascribed to disturbed secretion, etc. Undoubtedly the best method 
of determining the efficiency of the gastric motor mechanism is the ront- 
genological observation of the time necessary for this organ to empty itself 
entirely of the buttermilk-bismuth subnitrate or barium sulphate meal. 
The stomach should be entirely clear of such contents in 6 hours. 

In case of food the normal stomach should be empty in 7 hours no matter 
how large the meal. A common test of motility and a method of estimating 
the degree of insufficiency is as follows (Boas) : The patient, whose stomach 
has not been washed out, is given a simple evening meal but one of constant 
composit ion, as cold meat, bread and butter and tea. On the following 

15 Deutsch. med. Wochenshr., 1904, No. 15. 



366 CLINICAL DIAGNOSIS 

morning the stomach is thoroughly washed out. If food is found the motor 
insufficiency is marked (Grade B) . On the following afternoon the stomach 
is well lavaged, the same meal is repeated and on the next morning again 
washed. If the stomach is found empty the insufficiency is less than B 
(Grade C). If food is found it is more than B (Grade A). If the stomach 
contains food 7 hours after a full noon or morning meal, but none after a 
night's rest, the degree of insufficiency is least (Grade D). 

Ewald and Strauss have recommended that 1 spoonful of currant or 
raisin preserve be given with the evening meal. These seeds can be recog- 
nized the next morning in the stomach washings even though the patient 
has taken a large breakfast. 

If before breakfast the stomach contains over 100 c.c. of fluid motor 
insufficiency may be suspected and lavage will probably wash out food 
particles, but such a stomach will contain no fluid if it is washed out the 
evening before. 

The symptoms of extreme dilatation are those of the primary disease 
together with the vomiting of food eaten more than 7 hours, in some cases 
even 3 days, before. The repeated vomiting of food in the morning before 
breakfast is conclusive proof. But the symptoms of less marked disturb- 
ances of motor function are not as characteristic. The most constant of 
these is a discomfort in the epigastrium which begins about 3 hours after 
a meal when the expulsive movements begin, and which is relieved by soda 
or by a lunch. " Heart-burn " is evidence of slight retroperistalsis, the 
eructations of acid fluid of a slightly more marked grade, and nausea and 
vomiting of a still more severe retroperistalsis. 

For those patients who object strenuously to a stomach-tube and who have not 
access to a rontgenological institute, various chemical tests have been proposed. That 
of Ewald and Sievers is based on the belief that salol remains unchanged in the stomach 
but is split by the pancreatic juice and by bacteria to salicylic acid and phenol. The 
salicylic acid may easily be detected by the violet color of the urine when ferric chloride 
is added. This test assumes that the time of splitting and of the absorption of the salol 
and the time of excretion of the salicyluric acid remain constant, which is not quite true. 
One gramme of salol is given with the test breakfast and the urine is examined at inter- 
vals. The acid should appear in the urine at the latest in 75 minutes. But since the 
salol may enter the duodenum with the first portions of food as well as with the last and 
since, in cases of achylia, bacteria can split some of the salol (and mucus can also) Huber 
recommends that we test not alone the time of the appearance but also of the disappear- 
ance of the salicyluric acid, which should be in from 26 to 27 hours. The urine is there- 
fore examined 27 hours after the meal, and, if found, repeatedly at intervals of 3 hours. 

Sorensen and Brandenburg 16 recommend that we give the patient when the stomach 
is empty 300 to 500 c.c. of 3% protogen. One removes as much of this as possible in 
from y 2 to 1 hour. From 100 to 200 c.c. of water are then introduced and as much more 
as possible removed. The nitrogen in both fractions is determined by the Kjeldahi 
method and from this the amount in the stomach calculated. 

The composition of the gastric juice in dilated stomachs will depend 
on the disease causing the dilatation. In general there are 2 groups of cases, 
16 Arch. f. Verdauungskrankheiten, Bd. 2. 



THE VOMITUS AND GASTRIC CONTENTS 367 

one with acid and the other with anacid contents. The former includes 
cases of ulcer, the latter those of cancer and chronic gastritis. As a rule 
the mucosa of a dilated stomach becomes less sensitive to stimuli, owing 
perhaps to the constant presence of food and to the gradual development 
of a chronic gastritis, and so secretes less and less acid. 

Of 45 of our benign cases, 7 were hyperacid, 15 showed normal acidity and 9 were 
hypoacidity with, and 4 without, any free hydrochloric acid. 

By " hyperacidity " is meant the secretion of abnormally acid gastric 
juice during digestion, that is, while there is a normal stimulus for secretion. 
By " hypersecretion " is meant the secretion of an amount of gastric juice 
out of proportion to or in the absence of a physiological stimulus. With 
the exception of a few cases of nervous disease it is doubtful whether either 
occurs often enough to be important. 

Hyperacidity, Superacidity, Hyperchlorhydria, or Hyperaciditas Hydro- 
chlorica. — The secretion of a very acid gastric juice, that is, one containing 
an abnormal per cent, of free hydrochloric acid, has been assumed as the 
explanation of many gastric conditions with pyrosis, etc. In recent years 
the demonstration that gastric juice is normally" much more acid than was 
supposed, that extreme acidity need give rise to no symptoms, that the 
heart-burn, acid eructations, the vomiting of acid gastric juice, etc., are 
due usually to pyloric spasm and to slight motor insufficiency, and, finally, 
that the chemical analysis of the fluids which symptoms would indicate 
to be very acid usually are not, have thrown considerable doubt on the 
importance of hyperacidity as such (see page 349). 

Hyperacidity formerly was attributed to diseases of the central nervous 
system, to a defective nervous control of secretion and to changes in the 
mucosa. This term was used of cases with an acidity per cent, over 70. 
Often it is 100, sometimes it is 150 to 160 or more and the free HC1 from 
60 to 80. Following the test meal the total is over 70, even 100 or more, 
and the free from 50 to 60. Others (Meunier) say that the acidity alone 
is not important, but that the specific gravity of the contents must be low, 
from 1.007 to 1. 01 9 instead of from 1.022 to 1.040, as normally. 

The majority of the cases formerly termed hyperchlorhydria are doubt- 
less cases of duodenal ulcer, of gall-stones or of subacute appendicitis. 
Nevertheless some are secretory neuroses. For this last diagnosis the 
acidity should be very variable and vary definitely with the nervous symp- 
toms, an important point in a diagnosis at best difficult. 

Hypersecretion, Supersecretion, Continuous Secretion, " Gastrosuccor- 
rhea." — In cases of continuous secretion the formation of gastric juice is 
supposed to continue when the stomach contains no food, which would 
suggest as cause a disproportion between stimulus and response. The 
free acidity in such cases is relatively high, which is a valuable point in 
ruling out cases of motor insufficiency. Continuous secretion is proven 
by finding much acid gastric juice in a previously well washed fasting stom- 



368 CLINICAL DIAGNOSIS 

ach. This condition may be constant or intermittent. It may also be 
reflex as from a duodenal ulcer; a part of a general neurosis; a secretory 
neurosis, or the result of organic nervous disease. Among the last may be 
mentioned the gastric crises of tabes dorsalis. Among the more constant 
cases are those of gastroxynsis (Rossbach) and those seen in neurasthenia 
and hysteria, in myelitis, in general paralysis and even after the excessive 
use of tobacco. In the periodic or intermittent cases of Reichmann's 
disease, the patient's digestion during the intervals may be perfectly normal, 
then there occur sudden pains, acid eructations and the vomiting of a 
cloudy yellowish fluid, which at first contains food, then is pure gastric 
juice measuring often several hundred cubic centimeters in amount, the 
acidity of which may be normal or increased (the latter usually only when 
food is present). 

The chief symptoms of the continuous cases are a feeling of discomfort 
and of weight in the epigastrium, eructations of acid fluid and pain which, 
begins about an hour after eating and increases until the stomach has 
emptied itself into the duodenum or until the patient has vomited. Vomit- 
ing about midnight is a very characteristic symptom. The patient usually 
vomits large amounts (from 500 c.c. to 1000 c.c. or much more) of a thin 
fluid the acidity of which is normal or slightly above the normal. There is 
often pain before meals, which is relieved by eating. 

Many doubt whether continuous hypersecretion is ever " functional." 
They claim that it is always caused by some gross stimulus to secretion, 
as ulcer, stenosis of the pylorus, or by some condition favoring the retention 
of particles of food. 17 Foster and Lambert ls have shown that pyloric 
stenosis is a sufficient cause of continuous secretion; that the presence of 
retained food particles is not necessary to explain it. Other cases called 
nervous are supposed to be due to reflex disturbances from the intestine 
and are relieved by treating this organ. 19 

To diagnosticate chronic hypersecretion, Riegel recommends the fol- 
lowing routine. The stomach is emptied after a full meal at the height of 
digestion and its contents measured. The next step is to pass the tube 
some morning after a night during which the patient has eaten and drunk 
nothing and note the quantity of clear fluid obtained. Over 100 c.c. is 
considered pathological. Lastly, at evening the stomach is washed and 
finally emptied (very carefully, since it is hard to wash out all the food) 
and the contents obtained the following morning noted. 

A case reported by Thayer will serve as a good illustration. The symptoms were of 
2 years' duration ; the total acidity after the Ewald breakfast was 113; the fasting stomach 
always contained even 420 c.c. of acid fluid, the acidity per cent, of which often was 117. 
Digestion was good. 

17 Kaufmann, Am. Jour. Med. Sci., 1904, vol. cxxvii. 

18 Jour, of Exp. Med., 1908, vol. x., p. 820. 

19 Faber, Arch, f . Verdauungskrankheiten, Bd. 7. 



THE VOMITUS AND GASTRIC CONTENTS 369 

Nervous Dyspepsia. — Hyperacidity, hypersecretion, anacidity, etc., 
acid eructations, gas and sensations of all descriptions, some very painful, 
are symptoms of conditions which may be due to organic changes of the 
mucosa, to functional disturbances following bad habits of eating, poor 
food, etc., to reflex stimulation from gall-bladder or appendix or be a part 
of a general neurosis, the "nervous dyspepsia" so common in this country 
that the stomach specialists abroad speak of it as the "American disease." 
It is exceedingly difficult to separate the element due to food, rapid eating, 
disease, etc., from the reflex and the neurotic elements and in the majority 
of cases perhaps 2 or 3 coexist. The nervous dyspeptic usually has some 
organic reason for his gastric distress although a neurasthenic will often 
worry his normal subliminal gastric sensations into the sphere of conscious- 
ness. He is nervous and he has gastric symptoms. It is, however, a great 
mistake to consider the latter entirely a nervous trouble for the majority 
have a lesion which demands treatment. Among these are inflammatory 
conditions of the nose and nasal sinuses, tonsillitis, pyorrhea alveolaris, 
stomatitis, chronic glossitis, etc. Considering the amount of pus which 
these patients swallow we may well admire the resistance of the stomach. 
Among other causes are ulcer of stomach or duodenum and disease of the 
gall-bladder or appendix. 

Some of these " neurasthenics," have hyperacidity; more, slight sub- 
acidity; and many, normal acidity. Their subjective symptoms bear very 
little relation to the condition of the gastric juice. A patient with normal 
gastric juice or with hyperacidity may describe sensations quite similar 
to those of an anacid case, except perhaps that the hyperacid case is more 
apt to vomit than either of the other two. 

In the Johns Hopkins clinic we studied the records of 300 such cases admitted during 
4 years. In 82 there was hyperacidity and in 20 others the clinical features were those 
of hyperacidity although the total acidity was not over 70. Subacidity (total acidity 
less than 40) was present in 170 cases, in 61 of whom there was no free hydrochloric acid, 
and in 4 of these the fluid was practically neutral to litmus. 

In conclusion, we would urge that the diagnosis " nervous dyspepsia " 
be made only of cases definitely neurotic whose gastric function is usually 
quite normal and whose periods of dyspepsia would seem directly related 
to neurotic states. The great majority of cases of so-called nervous dyspep- 
sia are true dyspepsias of nervous persons who have definite organic basis 
for some at least of their troubles. 

Acute Gastritis. — By acute gastritis is meant an acute irritation or 
definite inflammation of the gastric mucosa, resulting, in mild cases, in 
increased mucus secretion and in more severe cases in an acute inflamma- 
tory reaction with desquamation of the epithelium, and in all with some 
disturbance of secretion. It may be due primarily to the direct irritation 
of unsuitable foods, to poisons of all sorts, or to heat, cold, etc. ; or, it may 
be a part of an acute infectious disease. The vomitus of these cases is 
24 



370 CLINICAL DIAGNOSIS 

acid, rarely neutral, in reaction, has a bad odor, is often fermented, the 
food is undigested as a rule and mixed with much mucus. The total acidity 
is diminished, free hydrochloric acid is absent as a rule and organic acid is 
often present. If there has been much retching the vomitus contains more 
or less bile. A test meal will show mucus, undigested food and little or no 
free HC1. 

We have records of but 5 clear cases, all of whom had subacid or neutral gastric 
contents. 

Gastritis phlegmonosa or interstitial purulent gastritis is a 
very rare condition with inflammation of the entire gastric wall even to the 
serosa. When localized a gastric abscess is the result. Vomiting is a com- 
mon symptom. In the 60 diffuse cases reported pus was not found in the 
vomitus (Riegel). It was found in a very few reported cases of abscess of 
the stomach wall. 

In the gastritis acuta purulentia (Leube) the inflammation is limited 
to the mucosa. 

Chronic Gastritis. — Chronic gastritis is not nearly as common a condi- 
tion as one would imagine from the number of times this diagnosis is made. 
The term implies a definite long-standing inflammation of the mucosa 
which leads to definite atrophic changes of this membrane and to weakening 
of the muscularis. It exists in all grades even to complete atrophy of the 
mucosa. Among the common symptoms are a great increase of mucus in 
the stomach washings; vomiting, especially on an empty stomach in the 
morning or at the height of digestion, of undigested or poorly digested food 
mixed with mucus ; and motor insufficiency . These cases have been classi- 
fied as primary, i.e., without demonstrable disease as adequate cause 
(although more and more the chronic pyogenic infections of the nose, 
throat, tonsils and teeth are considered satisfactory explanation) and 
secondary, or accompanying other diseases as ulcer or cancer of the stomach 
or pyloric stenosis from any cause whatever. 

The amount of gastric contents removed after a test meal is about 
normal. The food has the appearance as if it were just swallowed, except 
that it is intimately mixed with much mucus which renders its removal 
through the tube difficult and its filtration tedious. The amount of mucus 
present is best estimated by macroscopic examination. The stomach must 
be washed thoroughly and repeatedly since the most of the mucus appears 
in the later washings. 

In chronic gastritis the secretion of gastric juice diminishes progres- 
sively as the case advances until the achylia may be complete. The total 
acidity varies much and at times some of the acid may be free, hence a 
correct diagnosis requires repeated examinations. The secretion of hydro- 
chloric acid decreases first ; that of the ferments second (Bouveret believes 
that the easiest way to follow the progress of a case is to observe the rennin 
secretion) ; that of water next ; while during all this time the secretion of 



THE VOMITUS AND GASTRIC CONTENTS 371 

mucus may even increase. Proteid digestion is much impaired, that of 
carbohydrates not at all. The stomach often becomes somewhat (seldom 
much) dilated since its walls are weakened. The pylorus may be slightly 
obstructed by inflammatory thickening of its mucosa and slight motor 
impairment result allowing fermentation, yet seldom will more than a 
trace of organic acid be found in the contents. The presence of much 
undigested food in the stomach washings does not necessarily indicate 
stasis since it is a function of the stomach to retain food until it is digested. 
In many cases, however, the gastric motility is normal or even increased 
and symptoms of stomach trouble absent since the intestine vicariously 
will fulfill the gastric function. In cases of chronic gastritis the mucosa 
of the stomach is abnormally fragile and one often finds fragments of 
mucous membrane in the stomach washings. 

Some cases begin with an actual increase of the gastric acidity. The 
fasting stomach of these cases contains mucus enclosing many cell nuclei 
and an hyperacid juice. The reason these cases of gastritis acida would 
seem to be rare may be that this stage has passed before the patient consults 
a physician. 

Of 27 of the Johns Hopkins Hospital cases, 1 was slightly hyperacid 
(72 total acidity), in 10 the acidity was within normal limits and in 15 it 
was below 40. Nine of these 40 had no free acid. Four could be termed 
atrophic catarrh. In 1 case only 1 c.c. of bile-stained mucus could be ob- 
tained by the stomach-tube and at autopsy was found cirrhosis of the 
stomach wall. 

Mucus. — A little mucus is usually demonstrable in the contents of 
normal stomachs especially towards the end of the digestion period. Mucus 
is present in excess under the following circumstances : if the diet is par- 
ticularly rich in starch; in conditions of subacidity and anacidity of the 
gastric juice, though in these cases the increase is sometimes only apparent, 
for, while the normal amount may have been secreted an amount less than 
normal may have been digested; if the stomach contents are irritating to 
the mucosa, which seems to protect itself in this way; rarely, in cases of 
gastric hyperacidity, and these all may belong in the preceding group; 
and finally in all forms of chronic gastritis (" gastric catarrh "), both the 
primary form and that which accompanies cancer of the stomach, pyloric 
stenosis, etc. The largest amounts of mucus are found in cases of develop- 
ing achylia. It would appear that the secretion of mucus increases as that 
of the gastric juice decreases. A correct idea of the amount of mucus 
present in the stomach can be gained only by repeatedly filling and empty- 
ing the stomach, or, better still, by the use of a needle douche-tube, since 
it sticks tenaciously to the gastric wall. 

Mucus from the stomach is present as delicate transparent flakes, 
which are mixed with the food, which sink in water, and which contain the 
nuclei of leucocytes. Mucus from the air passages is present in glassy balls, 



372 CLINICAL DIAGNOSIS 

which float (they enclose air) , which contain cylindrical epithelial cells and 
often visible pigment and which are not mixed with the food. Foster 20 
attributes to the mucus the diminution of, and the variations in, the amount 
of free HC1 in cases with a high free acidity, for the products of the diges- 
tion of mucus have a high acid-binding power. 

Atrophy of the Mucosa — Achylia Gastrica. — Achylia gastrica may be 
due to a functional disturbance of an apparently normal mucosa or to real 
atrophy of this membrane. The latter may be the end stage of a chronic 
gastritis or accompany cancer and other diseases which lead to degenerative 
changes of this mucosa. One must remember that the diagnosis of achylia 
very often is an error since the meal was not removed at the proper time 
(see page 354). 

Some cases of achylia are of nervous nature, and the discovery purely 
accidental. In other cases the nervous symptoms may disappear but the 
achylia continue. When due to atrophy the process is gradual and the 
secretion may diminish until finally there is almost no gastric juice. There 
is an interesting group of cases of achylia associated with very small gastric 
cancers which is apparently toxic in character since much of the mucosa 
looks normal. Achylia gastrica sometimes accompanies cancer in other 
organs (breast, intestine, esophagus, uterus, etc.) and is present before any 
disturbance of the general health is manifest and may be due to other condi- 
tions which lead to general malnutrition. The cases of particular interest 
are those which simulate pernicious anemia and those with protozoon 
infection of the bowels. 

A severe achylia may not be suspected provided the motility of the 
stomach is good (the intestine seems to act vicariously) although if 
even slight motor insufficiency has developed the symptoms will be 
evident enough. 

An atrophied mucosa is very susceptible to injury. It is indeed not 
uncommon to find in the stomach washings pieces of diseased mucosa. 
Such cases often vomit undigested food soon after eating, which seldom 
contains blood. The diagnosis of achylia due to atrophy is made with the 
Rehfuss stomach-tube, testing the gastric contents at frequent intervals 
until the stomach is empty. The food removed is little changed. The 
total acidity is very low, from 1 to 4, there is no free HC1 and lactic acid 
is rarely present except in cases of severe ectasis. The ferments may be 
absent, an important point in diagnosis. In most cases the washings with 
the larger tube contain great quantities of mucus for the mucosa may 
consist chiefly of mucus cells; the glandular cells have practically disap- 
peared. In extreme cases even mucus is absent. To obtain nothing through 
the tube by simple siphon action does not necessarily mean that the 
stomach is empty and clean, since subsequent lavage may remove dry 
food. All of these findings should be confirmed by several test meals. 

20 Am. J. of Med. Sci., Feb., 1907, vol. cxxxiii; and Med. Record, Aug. 13, 1910. 



THE VOMITUS AND GASTRIC CONTENTS 373 

Ulcer of the Stomach and Duodenum. — In general the clinical picture 
is the same whether the ulcer is on the gastric or duodenal side of the 
pylorus. The clinical types of this disease are: (i) The latent ulcer which 
may pass unsuspected unless hemorrhage or perforation occurs. This is 
seldom near the pylorus or in the duodenum. 

(2) The hemorrhagic form, which may be acute and sometimes fatal, 
or chronic, the frequent small hemorrhages causing considerable anemia 
and cachexia. The stools of these patients constantly contain a certain 
amount of blood. 

(3) The acute perforative type. 

(4) The chronic dyspeptic type in which the symptoms of " indiges- 
tion " may persist for years. These ulcers are usually near the pyloric ring. 

(5) The neurotic, or gastralgic type. 

(6) The vomitive form, with vomiting the worst symptom. 

(7) The cachectic form which presents the picture of a cancer. 

The cardinal text-book symptoms of gastric ulcer are: (1) Gastric 
symptoms usually of long duration and beginning as a rule in young adult 
life. (2) Pain, paroxysmal and local, from half an hour to 2 hours after the 
meal, i.e., when peristalsis is most active. (3) Vomiting of acid vomitus 
containing well-digested food usually 1 to 3 hours after the meal but also 
often in the morning, and followed at once by a diminution of the pain. 
Hemorrhage, in from 30 to 50% of the cases and in these only occasionally. 
This blood as a rule is dark in color (due to the hematin formed by the 
hydrochloric acid) although if the stomach is empty at the time of the 
hemorrhage it may be arterial. There is often occult blood in the stools 
but it is not present as constantly as in cancer. If the blood remained for 
some time in the stomach before it was vomited or removed through a tube 
it may resemble coffee grounds. In such cases iron, wine, coffee, medicines 
and certain foods must be excluded. Other reasons for hemorrhage must 
be excluded, as tuberculosis, cancer, chronic passive congestion, cirrhosis 
of the liver, rupture of an esophageal varix, etc. (5) Hyperacidity is a 
classical symptom but the subjective sensations of this (pyrosis, acid eruc- 
tations, pain, etc.) often do not coincide with the chemical findings and 
•are due more to slight retroperistalsis than to hyperacidity. Ewald, using 
the Riegel meal, found in 75 cases that the total acidity averaged 105 and 
the free HC1 50, with a maximum of 89. But these figures are not very 
high and in at least half the cases the acidity is diminished. Many separate 
2 groups, the " fresh " and the "old " ulcers, and find that in the latter 
the acidity is much lower. Repeated examinations should be made in order 
to get a correct idea of the acidity conditions. 

In general in ulcer cases digestion is good and motility is rapid. 

The results of a gastric ulcer may be stricture of the pylorus or severe 
anemia due to the insufficient nutrition, vomiting and hemorrhages. In 
case the blood is lost chiefly by the stools, as in duodenal cases, the ulcer 



374 CLINICAL DIAGNOSIS 

may be unsuspected and the case be diagnosed pernicious anemia. It is 
said that in case a cancer develops on the bed of an ulcer the acidity may 
for a long time not be lessened, but in most cases it gradually diminishes. 

The 82 cases of the Johns Hopkins Hospital medical clinic were reported 
by Howard. 21 Vomiting was present in 85.3%, especially in the cases with 
ulcer at the pylorus. Blood was vomited in 75.6% of the cases although 
in % only was the blood bright red. After the test breakfast more than 
50 c.c. was obtained in 54% of the cases. Hyperacidity was present in 
2 7-5%> i n 4 2 -5% subacidity (for these figures an acidity per cent, of 60 
was considered the upper limit of normal). In 18% there was no free 
HO, while in 14% lactic acid was present. 

The laboratory tests of duodenal ulcers are as unsatisfactory as the clin- 
ical evidence is easy. Hunger pains entirely relieved by eating, ' ' heart-burn, ' - 
acid eructations or vomiting from 1 to 5 hours after a meal relieved by 
further eating or by soda, together with the intermittent presence of blood 
in the stools are enough for diagnosis. 

Hemorrhagic Erosions. — It is doubtful whether there is any symptom 
complex of hemorrhagic gastric erosions which is characteristic. The most 
suggestive evidence would be fragments of mucosa without marked patho- 
logical changes and found in the washings of the empty stomach. Vomiting 
is rare. The acidity in- such cases is normal or diminished, rarely increased. 

Syphilis of the Stomach. — Lyon 22 writes that the diagnosis of syphilis 
of the stomach is justified if the serological examinations are definitely 
positive and if syphilitic therapy results not only in a general clinical 
improvement but also in a cessation of gastric symptoms. 

This is hardly true since gastric symptoms may be evidence of disturbances not in 
the stomach but in other organs, as the gall-bladder, appendix, etc., and in luetic patients 
these may be relieved by antisyphilitic treatment, thus relieving the gastric symptoms. 

Being a tertiary lesion gastric lues is far more apt to become manifest 
during the middle decades of life, it may affect both sexes and may be a 
result of both congenital and acquired infection. 

Pathologically the disease may show any one of the following forms: 
(1) A diffuse gastritis; (2) syphilitic ulcers ; (3) a diffuse infiltration of the 
gastric wall; (4) pyloric stenosis and (5) gumma, which may or may not 
give rise to a palpable tumor. 

The secretory defect is usually accompanied by the symptoms of a 
severe atrophic or sclerosing gastritis. 

The gastric analysis shows a marked subacidity or anacidity with a 
greatly diminished or absent enzyme activity. On the other hand a few 
cases have been reported in which the hydrochloric acid content and peptic 
activity were normal or even increased. An increase of endogenous mucus 
is generally the rule. Occult bleeding is frequently encountered both in 
the gastric filtrate and in the feces. 

21 Am. Jour. Med. Sci., December, 1904. 

22 The Arch, of Diag., Apr., 1917. 



THE VOMITUS AND GASTRIC CONTENTS 375 

Cancer of the Stomach. — Clinically cases of cancer of the stomach may 
be separated into 3 groups : the latent ; those with cachexia but no gastric 
symptoms; and those with definitely localizing symptoms. The important 
points in the diagnosis of a quite advanced and typical case of this disease 
are: a history of the rather sudden appearance of dyspeptic symptoms in 
a person beyond 40 years of age, which symptoms were new to that person; 
loss of weight and strength ; anemia ; the lack of free hydrochloric acid and 
the presence of lactic acid in the gastric contents. But " typical " cases 
are rare. The sudden and apparently inexplicable loss of weight and 
strength and the development of a secondary anemia in an adult should 
always arouse our suspicion of malignant disease and yet such a case could 
not be called " early." It probably would be far advanced but so situated 
that its presence as a tumor could give no localized symptoms. It might 
be anywhere in the body. The additional symptoms which would indicate 
that the cancer involves the stomach may be divided into 2 groups: Those 
depending on the fact that the stomach is the organ involved and those 
which depend on the location of the cancer in the stomach wall. 

Undoubtedly the earliest and most important symptoms of gastric 
cancer are: a change in the appetite, especially a loss of appetite for meat, 
slight dyspeptic symptoms such as gas, a heavy feeling after eating, indefi- 
nite discomforts, etc., which are new to that patient and which have not been 
present for more than 2 years. Among the localizing physical signs are : 
a palpable mass and stenosis at cardiac orifice or at the pylorus. These 
differ in no way from those due to benign conditions. 

Among the early clinical laboratory tests are the almost continuous 
presence of occult blood in the stomach washings and in the stools and 
the irregularity in the curve of the free HC1. 

At present the rontgenological examination of the stomach is over-popular 
as a means of early diagnosis of this disease, one reason for which is, we firmly 
believe, that doctors have not been careful in taking case histories or 
accurate in their clinical laboratory work. The latter 2 methods are, we are 
confident, of far greater value in early diagnosis than is generally believed. 

Of the later local symptoms and signs, any one of which may be absent, 
are: vomiting, hemorrhage, subacidity and fermentation. Vomiting is 
very common (in 85.3% of Osier's first 150 cases), but it depends much 
on the position of the cancer. If it is at the pylorus causing stenosis the 
vomiting will be late after a meal but if at the cardiac orifice there will 
be regurgitation at once after eating. There is least vomiting when the 
cancer is on the lesser curvature. If there is no stenosis at either orifice 
there often is no vomiting at all, but it is also true that in 6 of 30 cases 
with the cancer actually at an orifice there were none. In late cases with 
dilated stomachs the patients may vomit from V 2 to 1 liter or more and in 
this may be recognized food that was eaten days (in 1 case four weeks 23 ) 

23 Osier and McCrae, Modern Medicine. 



376 CLINICAL DIAGNOSIS 

before. The proteins of the food will be found poorly digested, the meat, 
in lumps often covered with mucus, sometimes decomposed and often mixed 
with digested blood. In other cases the motility is excellent or even 
increased, as in n of 76 cases in which it was hard to get any fluid at all 
at the end of an hour. 

Some hemorrhage is almost constant in gastric cancer. It is parenchym- 
atous as a rule and yet early it may be profuse and even fatal. As a rule, 
however, the patient knows nothing of it unless his stomach be washed 
out or his stools are carefully examined, since it is gradual, and, since the 
blood is digested, it resembles coffee-grounds and is mixed with food. Of 
the 150 cases reported by Osier and McCrae, the vomiting of blood occurred 
but in 21.8%, but chemical examination of the stools shows it present in, 
almost 100% of the cases. 

Other symptoms, e.g., the gradual diminution in the amount of secretion, 
are due to the chronic degenerative changes of the mucosa which begin 
early and develop late. The absence of free HO is an early and important 
sign of cancer, present in over 80% of all cases on first examination, 
although there is one group without previous symptoms of ulcer which begin 
with hyperacidity (Zeigler). In 163 cases of Osier's clinic the free acid 
was absent in 146, or 89%. Alone, a subacidity is not of great importance 
since in cases of pernicious anemia, of chronic gastritis and of cholecystitis 
the free HC1 may fail, but in cancer one finds a reduction of the free HC1 
when the total acidity is not diminished and while the total chlorides, are 
high. The free and total acidities vary considerably from, day to day. 
Even in the presence of free acid this may suggest carcinoma and in a 
recent early case was the point which led to operation. In this case the 
3 Ewald breakfasts given during 14 days had the following acidities: total 
i2i f 100, 37; and free HC1, 11, 16 and 10 respectively. In early cases the 
free acid may suddenly disappear and then following the excision of the 
cancer may return in a day or two. In cases of carcinoma of the duodenum 
or esophagus the disappearance of free hydrochloric acid can best be 
explained as due to a fluid from the tumor which flows into the stomach. 

The diminution or disappearance of free HC1 certainly is due at first 
to the binding of this acid by some body which itself does not react alkaline 
to litmus. The idea originally suggested by v. Velden that the cancer 
furnishes a secretion which binds the acid is now again advanced. What 
these bodies are which bind the free hydrochloric acid has been the subject 
of considerable investigation. Certainly the hydrochloric acid introduced 
into the stomach of a carcinoma case is soon neutralized (Stahelin) . They 
could be products of the digestion of proteins, albumoses, peptones and 
hexone bases. Reissner explained the disappearance of free acid in the 
presence of an undiminished or even increased total acidity on the theory 
that free alkalies are secreted by the tumor tissue. But if this were the 
case the chlorides thus formed would be neutral and the total acidity would 



THE VOMITUS AND GASTRIC CONTENTS 377 

be reduced. Work which we have done leads us to believe that the tumor 
furnishes the gastric contents with a proteolytic ferment the action of which 
produces an abundance of digestion products which can bind the acid and 
yet allow it thus combined to react as acid to litmus. Fisher 24 has confirmed 
this work by isolating from the contents of carcinomatous stomachs tyrosin, 
leucin, arginin and lysin. 

Later in the progress of the malignant disease there is a gradual reduc- 
tion in the amount of total HC1 secreted which continues until finally the 
gastric contents may react as alkaline to litmus. This is due to degenerative 
changes in the gastric mucosa. On the other hand the secretion of hydro- 
chloric acid may for a time increase as the result of regular lavage, proper 
dieting and general building-up treatment. In cases of cancer developing 
on the base of an ulcer the presence of free HO may continue for a con- 
siderable period. In such cases there are often marked fluctuations in the 
quantity of acid. 

Of 64 of our cases with no free HC1 the gastric contents after the Ewald 
breakfast was almost or quite neutral in 8, below 10 (acidity per cent.) 
in 20, between 10 and 20 in 15, between 20 and 50 in 14 and between 60 
and 103 in 7. The higher acidities depended on the lactic (and butyric) 
acid present. 

Later the pepsin and rennin are diminished. This is due to the chronic 
gastritis and is not specific for cancer. 

The presence of lactic acid in the gastric contents is often an early 
and very valuable sign of cancer. It may be demonstrated in about 90% 
of all cases when none of the HC1 is free although the bound acid may be 
abundant. It is true that lactic acid cannot be demonstrated in all cases 
of cancer and also that it may be found in some benign conditions, as in 
cases with atonic and anacid stomachs and in atrophic catarrh with stenosis 
of the pylorus. But in the benign cases it is usually absent, even though 
the stenosis is extreme and the fluid anacid, so that its early appearance in 
cancer before the stenosis is marked and the total hydrochloric acid much 
diminished must indicate for it a different significance and emphasizes 
the value given it by Boas in the early diagnosis of malignant disease, 
although the specificity he claimed is no longer granted. 

Of 609 benign gastric cases lactic acid was present in 30. All were cases of subacidity 
with no free hydrochloric acid. " These cases were: atrophy of mucosa, 1; chronic gas- 
tritis with dilated stomach, 4; ulcer, 6; " nervous dyspepsia," with anacidity, 3; perni- 
cious anemia, 2; gall-stones, 1 ; cirrhosis of liver with jaundice, I ; cancer of gall-bladder, 
1 ; pulmonary and peritoneal tuberculosis, 3 ; an interesting group of inflammation of the 
large intestine (ulcerative colitis, etc.), 5; cancer of the ovary, 1; peripheral neuritis, 1 
and fibrinous pericarditis, 1. 

One might expect to find lactic acid in the gastric contents in cases 
with diminished secretion involving the ferments as well as the HC1, with 

24 Deut. Arch. f. klin. Med., April 24, 1908, Bd. 93, p. 98. 



378 CLINICAL DIAGNOSIS 

stagnation and perhaps with insufficient absorption (Hammerschlag) , yet 
Riegel found considerable in the gastric juice of cases of cancer without 
much atony and reported that as the result of his experience of over 20 
years the presence of considerable lactic acid practically always means 
gastric carcinoma. He explained the lactic acid in 1 case without stasis 
as due to acid-producing organisms retained in the fissures at the base of 
the cancer. 

The usual method to test for lactic acid is to examine the gastric con- 
tents removed 1 hour after an Ewald breakfast at the same time that one 
tests for HC1, but this is hardly wise since its formation cannot be as rapid 
as that of the secreted acid. It is better to wash the stomach out well the 
evening before and test the contents of the fasting stomach the next morn- 
ing and then again after a test meal which is free from this acid or bodies 
which could easily form it. 

Lactic acid may be formed by the organisms in the stomach, several 
of which have been proven to be acid-producers, among them the Boas 
bacillus; or it may be a normal product of digestion but found when its 
absorption is diminished (an improbable explanation in many cases); or 
it may be the product of a specific ferment furnished by the tumor. This 
is possible since in the autolytic digestion of proteid by ferments from these 
tumors lactic acid has been shown to arise. 

In Osier's cases the fluid removed. 1 hour after an Ewald breakfast was examined 
for lactic acid, hence the per cent, of its incidence will be minimal. Yet it was present 
in 63% of 137 cases. The figure given by Schiff was 73.5% of a group collected from 
various writers. 

The gross appearance of the gastric contents in cancer cases is of great 
importance in diagnosis since the meat is poorly and the carbohydrates are 
well digested. Disturbance of motility is due to mechanical obstruction 
at the pylorus. On the other hand the motility may be excellent and yet 
the digestion very poor, which is true of some early cancers not at the 
pylorus. Strauss claims that this abnormal fermentation is due to bacteria 
which remain in the clefts of the tumors and so are not removed by lavage. 

Tumor fragments are seldom found in the stomach washings. These 
should be looked for in all bloody masses. Sahli suggests that the stomach 
be well washed out at night and the fasting stomach again the next morning 
and this latter wash- water carefully examined. The fragments of mucosa 
frequently found in the wash water in cases of achylia gastrica may closely 
resemble fragments of cancer. 

Sarcinse and yeasts are rare in the contents of cancerous stomachs and 
their presence in large numbers is evidence against cancer. In but 5 of 
Osier's cases were sarcinae found. The bacterial flora of the stomach is 
abundant. The organism which has attracted the most attention is the 
so-called Oppler-Boas bacillus, which is said to be met with in about 8o% 
of cancer cases (Rutinmever) , and in almost no other condition. The 



THE VOMITUS AND GASTRIC CONTENTS 379 

cultural characteristics of this organism are in dispute, probably since 
several pass under this title and cultures are seldom made. By Oppler- 
Boas bacillus we usually have in mind a long, coarse, thread-like bacillus, 
often in long chains which extend across the field of the microscope. In 
some cases they are present in such enormous numbers that they even 
fill the whole field. No spores are seen. The single bacilli are from 3 to iom 
long (6 to Sfi as a rule) and i/u broad. They have rounded ends and often 
are slightly bottle-shaped. Some are bent. They do not decolorize by 
Gram's method and are best seen in stained specimens. It is rather agreed 
not to report the Oppler-Boas bacilli as present unless such bacilli are pres- 
ent in large numbers. Some of the organisms thus named may be the gas 
bacillus. The so-called Oppler-Boas bacillus is not anaerobic nor will it 
grow well on ordinary media, but will luxuriantly if blood or its derivatives 
be added. This may explain its frequent presence in the cancer stomach 
in which condition alone blood is almost constantly present. In cancer 
cases also are the ulcerations and clefts which would hide these organisms 
during lavage. Here also is there a failure of ferment and of acid secretion 
and stagnation all of which factors Schmidt considers essential for the pres- 
ence of this organism. Kaufmann 25 claims that this organism cannot grow 
if there is 0.02% of free HC1 present but can well if the acidity is due to 
phosphates and lactic acid ; but others (Rosenheim) say it flourishes in the 
stomach in spite of free HC1. It coagulated milk. Kaufmann found this 
bacillus in 19 of 20 cancer cases, proved that it was a lactic acid builder, 
and found that it occurred in numbers proportional to the amount of lactic 
acid present. Most other lactic-acid-forming bacilli are smaller. Other 
bacilli of similar appearance have been cultivated 26 which adds to the 
confusion. In a recent case without extreme stasis the gross sediment of 
the stomach washings was almost entirely composed of masses of these 
bacilli. If stenosis certainly is present and these bacilli are absent, the 
evidence is against cancer (unless the stomach has been too well washed out) . 

The Hopkins series is hardly suitable for purposes of statistics concern - 
ing the presence of these organisms, for only the fluids removed 1 hour 
after an Ewald breakfast (and the stomach had been washed out the even- 
ing before) were examined. These organisms were found in only 38% of 
55 such cases. In 4 of these lactic acid seems not to have been present. 

Heichelheim 27 thinks that in the diagnosis of cancer clots of blood in 
the gastric contents are very important, especially if they contain many 
coarse bacilli and the fluid is without free HCL 

Pus is sometimes present in the gastric contents; in fact the largest 
amount of pus that we have seen in any gastric case was one of carcinoma. 

The resorption through the gastric mucosa is much disturbed and the 
KI test is almost al ways delayed. 

25 Centralbl. f. inn. Med., 1904, No. 4. 

26 See Schmidt, Wien. klin. Wochenschr., January 10, 1901. 

27 Zeitschr. f. klin. Med., 1904, vol. liii, p. 447. 



380 CLINICAL DIAGNOSIS 

Among other tests proposed for the early diagnosis of cancer of the 
stomach is the tryptophan test of Erdmann and Winternitz 2S which is not 
constant enough (Sigel in 2 of 15 cases; Glassner in 1 of 2) to be of 
great value and is positive in other conditions (ulcer, e.g.) ; yet its presence 
does help. 

The presence in the washings of a fasting stomach with good motility 
of over 0.5 p.m. of albumin (Esbach) is of some, but not great, value. To 
determine this the stomach is carefully washed out, then a few hours later 
the tube again introduced and all the fluid possible obtained. The stomach 
is then washed several times with 400 c.c. of physiological salt solution. The 
albumin and nitrogen of this gastric'fluid are then determined. In all other 
conditions N = o to 16 mgms. but in cancer from 10 to 70 mgms. per 100 c.c. : 
cancer is probable when N=more than 20 mgms. and there is a definite 
albumin precipitate by Esbach's fluid. This albumin is supposed to come 
from the ulcerations of the cancer nodule which furnished this inflamma- 
tory exudate. 

Gluzinski's test for the relative insufficiency of HC1 secretion was pro- 
posed especially to indicate a cancer on the bed of an old ulcer. He tested 
the free hydrochloric acid in the morning on the fasting stomach, 45 minutes 
after the test breakfast and again 4 hours after a full meal. The test is 
positive when the stomach fails to respond suitably to the greater stimulus. 

Infusoria and flagellates especially are sometimes present in very early 
stages in anacid carcinomatous stomachs. Cohnheim reported 6 cases 
with Trichomonas and Megastoma entericum present. He thinks this a 
valuable sign, even the earliest, for the diagnosis of an ulcerating cancer 
of the cardia or lesser curvature, but not of cancer of the pylorus, since the 
lactic acid present in these cases would kill them. They are often present 
in our food. Zabel 29 reported 4 early cases with such organisms present 
in abundance. Rosenfeld 30 found them in 6 cases, 1 of which he thinks 
is the first non-carcinomatous case in which they had been found. He 
thought this was true of still another case, but that proved to be a case of 
cancer. They are found in the small amount of neutral or alkaline fluid 
of these fasting stomachs, together with leptothrix threads, long bacilli 
and spirilla. It is interesting that they cannot be found in other cases of 
achy Ha for we certainly must swallow them frequently with our food. 

Blood, usually occult, in the gastric contents and stools is a very im- 
portant, a common (68 of 70 cases) and early sign of cancer, especially 
in the absence of free hydrochloric acid and when motility is good. 

Strauss emphasized the disproportion between the relatively active 
fermentation and small amount of sediment in case the cancer is not at the 
pylorus; Reissner, the early increase of chlorides to almost or quite the 

28 Munch, med. Wochenschr., 1904, p. 299. 

29 Wien. klin. Wochenschr., 1904. 

30 Deut. med. Wochenschr., 1904. 



THE VOMITUS AND GASTRIC CONTENTS 381 

double and the alkaline reaction of the ashed gastric contents. For Glass- 
ner's idea concerning ferments see page 356. 

The " early " diagnosis of cancer of the stomach is unfortunately usually 
a late one if one means by " early " a time when the patient can be saved 
by operation. No one feature will help. All evidence must be sought for 
and carefully evaluated. The chemical signs may be very suggestive in 
some cases, in others normal, while in some even the reverse of those sug- 
gesting cancer. The surgeons insist that the diagnosis should be made long 
before a tumor is palpable and this should be the aim of the clinical chemist. 
At present we feel that age and clinical history are more important than 
chemical examination and this more important than the rontgenological 
examination. We would never delay operation until the diagnosis was 
positive but would take a chance if a suggestive subjective history were 
strengthened by one objective sign, whether chemical, rontgenological 
or physical. 



CHAPTER IV 
THE INTESTINAL CONTENTS AND THE FECES 

Pancreatic Fluid. — The duodenal contents were formerfy obtained by 
massaging the contents of the duodenum back into the stomach which 
previously has been washed with a i% soda solution. The abdomen of 
the patient lying on his back was massaged from right to left from the 
costal margin to the parasternal line. The stomach -tube was then quickly 
introduced and all possible contents removed. Sometimes about 50 c.c. 
are obtained. To prevent the destruction of the ferments by hydrochloric 
acid, soda should be added at once. A much more satisfactory method 
however is made possible by Einhorn's duodenal tube. 

The presence of trypsin in the fluid obtained is assumed if fibrin or egg 
albumin is digested in alkaline medium. 

Trypsin — Arthus and Huber's Method. — Fresh fibrin is washed in 
water, then heated at 40 C. for 24 hours with 2% NaF and filtered. The 
intestinal fluid plus an equal amount of 2% NaF is mixed with 2 or 3 vol- 
umes of the above-described fibrin and kept for some time in the thermostat 
at 40 C. If trypsin is present the typical crystals of tyrosin will easily 
be found. Contamination with bacteria need not be feared. 

The fat-splitting ferment may be demonstrated as follows : Neutral 
olive oil is obtained by shaking out olive oil with ether and a little NaOH. 
The ether extract is shaken out repeatedly with water and the ether then 
evaporated. It is then emulsified by shaking together 10 parts of oil, 
5 of gum, and 35 of water. In each of 4 test-tubes are mixed 10 c.c. of very 
dilute neutral litmus solution and 5 drops of this emulsion. The fluid to 
be tested is added, 2, 4, 6 and 8 drops respectively to the 4 tubes, and 
these are left for a few minutes in a water-bath at 37 C. The presence 
of lipase will be indicated by the red color in one or more of the tubes. 

The presence of diastase is proved if at the end of a few minutes after 
a little of the fluid has been mixed with a thin starch solution the addition 
of a drop of iodine solution fails to give a blue color (see page 357). 

Brown's method of estimating the functional activity of the pancreas 
by estimating the diastase of the stools is as follows : l 

Brown estimated the diastase of the stools in preference to trypsin or lipase since 
both of these are easily destroyed by bacteria, since trypsin digestion may be simulated 
by the action of erepsin and by that of the putrefactive bacteria of the intestine and since 
trypsin must be activated by enterokinase and lipase by bile. On the other hand prac- 
tically all of the diastase in the bowel is secreted by the pancreas and being a preformed 
ferment does not require an activator. 

1 Johns Hopkins Hospital Bull., July, 19 14. 
382 



THE INTESTINAL CONTENTS AND THE FECES 383 

Diastase converts starch to maltose the intermediary products being soluble starch, 
erythrodextrin, achroodextrin and isomaltose. Since constipation lessens the amount 
of diastase while some purgatives {e.g., senna) increase it, Epsom salts was chosen as 
the best laxative. 

To exclude the salivary diastase liquid food is used to eliminate chewing. 

There are many sources of possible error in this method but the following routine 
will reduce them to a minimum. After a very light evening meal the patient at night 
is given a high enema. At 7 a.m. the next day he drinks 750 c.c. of milk, and at 7.30 
a.m. and again at 8 a.m. he is given half an ounce of Epsom salts. At 8.30 a.m. he 
drinks a glass of water containing % of a teaspoonful of bicarbonate of soda. All the 
stools up to 2 p.m. are saved in a vessel containing 2 ounces of toluol and kept on the 
ice or in a cool room. If less than 400 gms. or cubic centimeters of stool are obtained 
an enema of a pint of water is given since in the average case between 400 and 1 100 c.c. 
of stool may be expected. 

The stool should be examined as soon as possible. It is first diluted up to 3000 c.c. 
with normal salt solution, stirred until absolutely homogeneous, a portion centrifugal- 
ized for 5 minutes and the supernatant, fairly clear fluid used for the tests. 

Diminishing amounts of this fluid are put into a series of tubes, 1.8 c.c. in the first, 
1.6 c.c. in the second, 1.4 c.c. in the third, 1.2 c.c. in the fourth, 1 c.c. in the fifth, 0.8 c.c. 
in the sixth, 0.6 c.c. in the seventh, 0.4 c.c. in the eighth, 0.2 c.c. in the ninth, 0.1 c.c. 
in the tenth, 0.05 c.c. in the eleventh, and 0.025 c.c. in the twelfth. The fluid in each 
of the tubes is then brought up to 2 c.c. with normal salt solution. To each of the 
tubes are added 2 c.c. of 1% solution of soluble starch (Kahlbaum). The tubes are 
now incubated at 38 C. in a water bath for half an hour,then cooled by the addition of 
tap water to the bath or by placing them under the cool tap, and are then tested quickly 
with a few drops of 0.1N iodine solution. The limit is that tube before the one in which 
the first definite blue color appears. The exact figures would be somewhere between 
this and the next succeeding tube. Slight variations in the temperature of the water 
bath and in the reaction of the medium have so little influence on the result that it is 
not necessary to reduce all the specimens to the same reaction to litmus. If a previous 
test showed a negative result in the first tube, or if very low readings are suspected, 
a supplementary series of tubes containing respectively 2 c.c, 3 c.c, 4 c.c, and 5 c.c. of 
the centrifugalized mixture is used. The unit chosen is the digestion of 1 c.c of 1 % 
starch solution at 38 C. in half an hour. The lowest normal reading in the series of cases 
studied was the tube 10, which indicated 60,000 units; if 1 c.c. could digest 2 c.c. of 1 % 
starch solution then 3000 c.c. could digest 60,000. The highest was 240,000. 

Examination of Bile from the Duodenum. — Lyon 2 noting the obser- 
vation of Meltzer that magnesium sulphate in the duodenum produces a 
relaxation of the sphincter of the common bile duct, proposes the 
following method of obtaining the bile for the purpose of the diagnosis 
of hepatic conditions. 

The mouth of the fasting patient is rinsed thoroughly with any good 
antiseptic solution, preferably potassium permanganate, 1 grain to 2 ounces 
of water, followed by a rinse with a weak solution of zinc chloride. A sterile 
duodenal tube fitted with any one of the later modifications of the original 
metal tip, is passed into the stomach. The fasting gastric residuum is 
aspirated into a sterile vessel, the stomach thoroughly rinsed and the 
patient, lying on the right side with hips elevated, is given a glass of water 
to drink while slowly swallowing the tube to the duodenal mark. 

2 Jour. A. M. A., Sept. 27, 1919, vol. 73, p. 980. 



384 CLINICAL DIAGNOSIS 

The tube usually passes into the duodenum in from 15 to 45 minutes. 
(This can be told by the duodenal tug and the character of the fluid aspi- 
rated.) When certain that the tube is in the duodenum a barrelful of air 
is introduced from a 1 ounce capacity syringe to balloon out the duodenal 
walls from the metal tip (to prevent traumatism of the duodenal mucosa) , 
a connection is made with a sterile aspirating vacuum bottle and gentle 
aspiration of the duodenum is begun. 

The contents of the fasting duodenum should be bile free, pearly gray, 
of syrupy and stringy consistency, fairly transparent and should have a 
relatively small amount of flocculent or flaky sediment. In cases of duode- 
nitis the gross appearance, the microscopic sediment and the chemical tests 
of this fluid differ widely from the normal. 

The first bottle is now detached and from 50 to 100 c.c. of a sterile, 
25% saturated solution of magnesium sulphate is introduced by means of 
a sterile syringe or by the gravity method from a sterile container. The 
tube is attached to the second sterile bottle and aspiration gently started. 
In from 2 to 10 minutes usually bile is obtained staining light yellow the 
magnesium sulphate solution still in the duodenum. When the color 
deepens to a pronounced yellow the material already collected in the second 
bottle is decanted into a sterile glass container, the bottle is reattached and 
biliary drainage is continued. The first bile aspirated is, Lyon believes, 
the bile present in the bile ducts, probably only the common duct. It 
measures from 10 to 20 c.c. in amount, is lighter yellow in color, is more 
likely to be transparent and is much less mucoid than the bile seen later. 
Then the bile suddenly becomes darker, more viscid and more concentrated. 
In normal gall-bladders it remains transparent, but is more of a molasses- 
yellow. Lyon believes this bile to be that stored up in and delivered from 
the gall-bladder. This bile varies in amount from 30 to 100 c.c, in 1 case 
it was 166 c.c. Lyon believes that the high normal should not exceed 
75 c.c. When it appears the bottle is replaced by a third sterile collecting 
bottle into which the bile is allowed to flow until it is replaced by a lighter 
yellow, thinner and usually transparent bile which is aspirated much more 
slowly and intermittently. This, Lyon believes, is bile freshly secreted 
from the liver. When this second transition appears the third bottle is 
detached and a fourth sterile bottle attached to collect the liver bile. 

In this way Lyon believes we can collect separately bile from different 
parts of the biliary apparatus and, by receiving them in separate containers, 
can study each sample separately. This method has already proved itself 
of great value. The following are a few illustrations of the interesting 
results recently obtained by this method in the surgical wards of this 
hospital. 

Hosp. No. 10590 is a woman aged 36, admitted Nov. 17, 1920, with a clear history 
of cholelithiasis. She was admitted with pain in epigastrium, nausea, vomiting, jaundice, 
clay-colored stools and bile in the urine. The rontgenograms showed a distinct shadow 



THE INTESTINAL CONTENTS AND THE FECES 385 

in the gall-bladder region. The bile obtained from the gall-bladder by Lyon's method 
was definitely purulent. Following this i aspiration the pain was at once relieved, the 
jaundice cleared up rapidly and the patient insisted on her discharge Nov. 22, 1920. 

Hosp. No. 10455 was a woman aged 27 admitted Oct. 18, 1920, with a diagnosis of 
cholelithiasis. On aspirating the bile from the duodenum definite gall-sand was obtained. 
On operation later similar sand was found in the cystic duct. 

Hosp. No. 1037 1 was a boy 16 years old admitted Oct. 1, 1920, as a case of conva- 
lescent typhoid fever with symptoms of cholecystitis. His Widal on admission was 
positive and his blood culture negative. From the bile obtained by Lyon's method from 
gall-bladder a pure culture of Bacillus dysenterise of Shiga was obtained. 

The last case to mention was an out-patient, a woman 26 years old, examined Nov. 
6, 1920, for symptoms of cholecystitis. Pus was obtained from the gall-bladder by 
Lyon's method. In this pus numerous streptococci could be seen in smears but none 
grew in the culture which gave a pure growth of a diphtheroid organism. This 1 aspira- 
tion of bile gave the patient great relief from symptoms. 

Motility of the Intestines. — It often is important particularly in metab- 
olism experiments to determine the motility of the intestine in order to 
separate the stools belonging to a definite period and to determine the 
presence of a latent constipation. The normal time of passage of food 
from pylorus to rectum after a mixed meal is from 6 to 20 hours; after 
milk, from 36 to 48 hours.' The motility of the bowels may be determined 
by watching the progress of a barium sulphate meal by means of the fluoro- 
scope and yet this cannot wholly replace the use of charcoal, lycopodium 
powder, or 0.5 gm. of carmine. After giving any one of these substances 
the stools are watched until the black charcoal is seen grossly, or until 
the characteristic lycopodium spores are seen microscopically, or the red 
color appears. Allowance should be made for the gastric motility in case 
the actual time in the intestine is desired. Since the charcoal mixed in a 
considerable mass of feces may pass unnoticed, lycopodium or carmine 
are somewhat safer substances to use. 

Test Meals. — To study intestinal conditions one should use a test meal 
with which he has had considerable experience especially with healthy 
persons. Folin's diet should be used by those who wish to compare their 
results with his, and he is the only one who has published a complete analy- 
sis of the urine of patients on any standard diet. 

The patient's diet is as follows (Folin): whole milk, 500 c.c; cfeam (18 to 22% 
fat), 300 c.c; eggs (white and yolk), 450 gms.; Horlick's malted milk, 200 gms.; sugar, 
20 gms.; sodium chloride, 6 gms.; water, enough to bring the whole up to 2000 c.c; 
extra drinking water, 900 c.c This daily ration consists of about no gms. of protein, 
148 gms. of fat, and 225 gms. of carbohydrate. It contains 18.9 gms. of N., 5.9 gms. of 
P 2 5 , 3.8 gms. of SO 3, and 6.2 gms. of CI. 

Others prefer for experiments in metabolism a diet of milk alone or of 
milk and eggs. In following a single patient through several periods of 
observation any diet will do providing it is constant, but if different patients 
are to be compared a standard diet is necessary. If the work is to be at all 
worth while the actual food used must be analyzed. Tables of food compo- 
sition cannot be used. 
25 



386 CLINICAL DIAGNOSIS 

McCrudden's 3 method of analyzing food given in metabolism experiments is fret 
from many of the objectionable features of the usual methods. Samples of the foods 
given the patient are saved for analysis, are all mixed together, in the proportions in 
which they are given the patient, so that I analysis of the mixture as a whole is sufficient. 
In this way one avoids a separate analysis of each food consumed, the food need not 
get stale while the experiment is in progress, one can give fresh food and change the diet 
as often as desired. 

In the case of liquid foods the fluids are mixed as thoroughly as possible, a certain 
volume is given the patient, and the same volume is taken for analysis. A solid food 
is mixed after being cut into small pieces. The patient receives a certain weight of this 
and the same weight is reserved for analysis. Only nonhomogeneous foods are excluded. 

At the end of the experiment all of this food reserved for analysis is well mixed 
together, a little HC1 added to retain all the nitrogen, as much as possible of its water 
is removed by evaporation on the steam-bath and the remainder by adding alcohol 
twice and continuing the evaporation. The food thus dried is next ground up in a grinder 
and then, since it can more easily be reduced to a powder when free from fat, this is 
extracted with naphtha. It is now crushed fine, so that it will all pass through a fine 
sieve and then its volume is reduced by quartering. That is, the food is thoroughly 
mixed with a large spatula and then made into a little circular pile 2 inches high. This 
pile is divided into 4 equal quarters. Two opposite quarters are rejected, and the other 
2 are well mixed together, made into another little pile, and quartered again. The 
mixing and quartering are repeated until there is about enough food left for 1 set of chem- 
ical analyses. 

For the macroscopic and microscopic study of the stools with a view to 
determining how well the various foodstuffs are utilized, the best diet is 
that of Schmidt and Strassburger. 4 

Morning: 0.5 liters of milk and 50 gms. zwieback. Forenoon: 0.5 liters oatmeal 
gruel strained (made from 40 gms. oatmeal, 10 gms. butter, 200 gms. milk, 300 gms. 
water and 1 egg). Noon: 125 gms. chopped beef (raw weight) broiled rare with 20 gms. 
butter, 250 gms. potato broth (made of 190 gms. mashed potatoes, 100 gms. milk, 10 gms. 
butter). Afternoon: As morning. Evening: As forenoon. 

This daily diet, which is given for a period of 3 or4 days, consists of 1.5 liters milk, 
100 gms. zwieback, 2 eggs, 50 gms. butter, 125 gms. beef, 190 gms. potatoes, and 80 
gms. oatmeal. 

It contains about 102 gms. proteid, in gms. fat, 191 gms. carbohydrates. Its 
total caloric value is 2234. 

In cases of jejunal fistula it is often important to know how near the 
opening is to the pylorus. Cushing 5 tied a silk thread to an oyster which 
the patient then swallowed and which soon appeared at the orifice. By 
measuring the length of the string this was found to be but 1 foot below 
the pylorus. This patient drank a glass of milk which began to escape 
from the fistula in 1 minute and all of which was recovered in 4 minutes. 

The examination of the stools deserves much more attention than it 
receives. Just as the sputum examination is commonly limited to a search 
for the tubercle bacillus, so that of the feces is now a matter of searching 
for parasites' ova and much that is valuable passes undiscovered. 

3 Jour, of Med. Research, 1903, vol. ix, p. 135. 

4 Die Faeces des Menschen, Berlin, 1905. See also Hewes, Boston Med. and Surg. 
Jour., April, 1909, vol. clx, p. 429. 

5 Johns Hopkins Hosp. Bull., July, 1899. 



THE INTESTINAL CONTENTS AND THE FECES 387 

All that is necessary for this examination are a few tall glass jars in 
which the stools mixed with water are allowed to sediment, some strainers 
(colanders) of various sized mesh through which the stool may be ground 
by a pestle, some large centrifuge tubes and plates half black, half white, 
similar to those used in sputum examination. 

The normal stools consist of the undigested portion of food, bacteria, 
intestinal secretions, formed and unformed elements from the mucosa, 
salts and products of digestion. The amount per day varies widely with 
the diet, but a general average is from 120 to 250 gms. 

The bodies of bacteria, the most of them dead, make up about one-third 
of the entire weight of the dried stools, that is about 8 gms. per day, and 
contain about % the nitrogen of the stools. The stools of some dyspepsias 
contain more, even from 14 to 20 gms., and strange to say those of cases with 
chronic constipations less, from 2.6 to 5.5 gms. Strassburger's method of 
determining the weight of bacteria was as follows : Two cubic centimeters 
of the stool is well mixed with water and centrifugalized ; the organisms will 
remain suspended while the elements of the food will sediment. The fluid 
is then decanted, considerable alcohol added to lower its specific gravity, 
and it is again centrifugalized to sediment the bacteria. The sediment is 
then dried and weighed. Another 2 c.c. are then evaporated (fresh alcohol 
being repeatedly added), dried and weighed, to get the total weights of 
solids in 2 c.c. of stool. Klein 6 used a counting method and takes exception 
to Strassburger's method and results, but he does not even suggest a guess 
as to the relative weight of the bacteria in the stool. 

The small amount of feces during starvation periods consists of bacteria, 
the intestinal epithelium, mucus and the intestinal secretions. 

Reaction. — The normal stool is usually alkaline in reaction, but may 
be neutral or faintly acid for the changes on standing are very rapid. The 
reaction at the surface of a mass of feces may differ from that at the center. 
If it contains any urine it will of course soon be alkaline. The stools of 
typhoid or cholera patients are alkaline as a rule, those of patients on milk 
or starch diet may be very acid. 

Frequency. — By diarrhea is usually meant the passage of frequent and 
fluid stools; by constipation, infrequent movements of the bowels, associated 
with symptoms which are relieved by purging. The normal stool is never 
fluid but frequency is a variable matter and must be judged from the indi- 
vidual standpoint. Some patients are very uncomfortable unless they 
have 2 movements a day while others normally have but 1 each 2 days. 
Much more attention has lately been paid to the time required for food to 
pass through the bowel, for some very constipated persons may have a 
movement regularly every day (latent constipation) while others badly 
constipated have several movements a day, hence the old adage, that 
diarrhea is one of the best symptoms of constipation. In these cases the food 

6 Zeitschr. f. klin. Med., 1903, Bd. 48, p. 163. 



388 CLINICAL DIAGNOSIS 

collects as scybalous masses in the colon and these by their mechanical 
irritation and the resulting infections of the colon wall cause the passage 
of frequent fluid stools often containing fragments of the hard dry masses 
from above. 

Diarrhea may be due to increased peristalsis, increased intestinal 
secretion or decreased absorption, and accompanies chronic enteritis, 
intestinal tuberculosis, amyloid disease, cirrhosis of the -liver, cholera, 
typhoid, dysentery, infectious disease, uremia, etc., as well as various 
functional nervous disorders. 

When the trouble is in the small intestine the movements are fluid 
and large but not necessarily very frequent ; in dysentery they are frequent 
and scanty. 

Constipation as a chronic condition is the result : (a) of careless habits 
assisted by a sedentary life and a diet lacking in the constituents which 
stimulate intestinal peristalsis; (b) of dilated stomach, constriction of the 
bowel, chronic appendicitis and chronic cholecystitis, etc.; (c) of general 
muscular atony. Acute constipation occurs in obstruction of the intestine, 
paralysis of its wall as in peritonitis, meningitis and other conditions causing 
increased brain-pressure. In acute obstruction due to intussusception, 
ileus, etc., the frequent stools of bloody mucus which contain no fecal 
matter may deceive the doctor who does not personally inspect them. 

The consistency and form of the normal stool vary considerably, de- 
pending on the habit and diet of the individual, and pathologically on the 
intestinal secretion, absorption, and, especially, motility. The stool may 
be abnormally too fluid or too solid ; when very hard it breaks up into small 
masses resembling sheep manure, or somewhat larger masses, " scybala," 
which may be of the size of a walnut and of even stony hardness. Such 
stools are common after typhoid fever and in some cases on a milk diet. 
These masses may in the rectum form large accumulations. For the mass 
of the stool to be of very small caliber does not necessarily indicate a stric- 
ture of the rectum since stools of small caliber are common in conditions 
of anal tenesmus, of inanition and in various nervous diseases. Boas 
believes that homogeneous, thick, pasty or curd-like stools in which float 
cylinders of fecal matter about the size of the little ringer suggest stenosis 
of the lower bowel provided the most of the stools are of this character. 

The feces are abnormally soft if they contain an excess of water, fat, 
fruit or of vegetable matter. An abnormal water-content may mean too 
rapid peristalsis of the colon (which precludes the normal drying of the 
stool) or an abnormal secretion of water by the colon. For the occurrence 
of fatty stools and of stools with excessive mucus see pp. 387 and 406. The 
most common vegetable foods which make the stools soft are cabbage, 
pears, apples, plums, etc. An easy way to determine whether the softness 
of a stool is due to fat or to water is to press a cover-glass down hard on a 
small portion. If when the pressure is relieved the cover-glass springs 



THE INTESTINAL CONTENTS AND THE FECES 389 

back and air rushes in from all sides, there is an excess of water ; if it stays 
as pressed, the softness is due to fat. Frothy stools indicate intense fer- 
mentation. Such stools often appear acholic (see below). 

Color. — The dark color of the normal stool is due to hydrobilirubin. 
Except in the case of nursing children bilirubin itself is never normally 
present. The longer a stool remains in the bowel and the longer it is ex- 
posed to the air, the darker it is. Certain foods influence its color. Milk 
makes stools light; meat, dark; cocoa, reddish brown; wines, dark; foods 
containing chlorophyll, greenish. Several drugs also are important. Calo- 
mel sometimes makes stools green (biliverdin) ; bismuth subnitrite, black 
(bismuth suboxide); senna, santonin, gamboge and rhubarb, yellow; while 
the stool containing iron turns dark, even black, after it has stood for some 
time in the air. A stool which contains digested blood is dark when 
first evacuated. 

A clay-colored stool may owe its color to an excess of fat which masks 
its pigment, or it may be dilute (as in diarrhea), while others owe their 
lack of color to the action of the organisms of putrefaction which reduce 
the bile pigments to colorless derivations: but the most important reason 
for the clay color is the absence of bile in the intestine (as in obstructive 
jaundice). If the paleness is due to an excess of fat the stool may be ex- 
tracted with alcohol and ether and the presence of bile demonstrated; if 
it is due to putrefaction, the color will be restored by exposure to air and 
the passage of such stools will cease after a dose of calomel. Stools free 
from bile, the ''acholic" stools, are of a grayish white color, have a bad 
odor and contain much fat. 

Bilirubin during its passage through the small bowel is so completely 
reduced to hydrobilirubin that under normal conditions practically none 
(except possibly minute traces enclosed in vegetable or soapy masses) 
reaches the cecum or the ascending colon. The stools contain bilirubin 
however in cases of diarrhea due to a peristalsis so rapid that the usual 
reduction cannot occur, and the higher up in the bowel the disturbance 
begins the more bilirubin will there be in the stools. It will appear in 
large amounts in the stools, therefore, if this absorption is disturbed and 
intestinal peristalsis very rapid. This is the case in simple diarrhea. The 
derivatives of bilirubin may be greatly increased in the stool, even to 400 
mgms. in 24 hours, if an abnormal amount of bilirubin enters the bowel 
with the bile, as in family jaundice. 7 

Since there is considerable bilirubin in the stools in cases of simple 
colitis some claim that normally much bilirubin does reach the ileocecal 
valve, but otheis believe that the so-called colitis practically always is an 
ileocolitis, or at least that the motility of the ileum is disturbed by a colitis. 

Stools containing much bilirubin (and biliverdin) are intensely yellow 
or green. Sometimes an obstructive jaundice is suddenly relieved and the 

7 Tileston and Griffin, Am. Jour, of Med. Sci., June, 1910. 



390 



CLINICAL DIAGNOSIS 



next movement will be large, soft, of a deep golden yellow color and give 
an intense Gmelin reaction. In the great majority of cases, however, the 
presence of bilirubin in the stools is not grossly evident and must be deter- 
mined by the microscope. For this, Schmidt's test is recommended. 




Schmidt's Test. — About 2 or 3 c.c. of the fresh stool, consisting of particles selected 
to represent as many as possible of its diverse constituents, are covered in a porcelain 
dish with a saturated aqueous solution of HgCl 2 (only the pure salt" should be used) 
and are then ground fine with a pestle so that the mercuric bichloride will mix thoroughly 
with the stool. The reaction of the mixture should be acid. The dish is then covered 
and allowed to stand 24 hours at the end of which time the fragments are examined 
macroscopically and microscopically. The particles stained with bilirubin will have 
turned green, those with hydrobilirubin, red. The green masses containing chlorophyll 

must be excluded by mi- 
croscopic examination. 
In diagnosis the 
green strands of mucus 
are most important. If 
large, they are probably 
from the colon; if small 
probably from the 
small intestine if the 
stool is fluid and espe- 
cially if the mucus con- 
tains the nuclei of many 
cells the protoplasm of 
which is digested, or 
contains cells repre- 
sented by fat droplets 
or bilirubin granules. 
Bile-stained muscle- 
fibers, connective tissue 
masses and fat masses are not as convincing since these normally are stained with bili- 
rubin while in the small intestine, and their presence in the stool may mean merely 
increased peristalsis or catarrh. These would suggest trouble in the small intestine 
only if the masses of bile-stained mucus also are present. 



■Wh- :: Qb 
b 






i 



mpmm 



Fig. 72. — Forms of fats and soaps in stools (Schmidt and Strassburger). 
a, soaps; b, casein and fat globules; c, fatty acid needles and leucocytes; 
d, yellow calcium soap; e, fatty acid crystals projecting from fat drop- 
lets; /, fatty acid and soap needles and scales from an acholic stool. 



Bile acids normally are so completely absorbed from the bowel that 
none appear in the stools. 

Fatty Stools (Figs. 72 and 73). — If the food contains much fat there 
always will be some in the stools, either in the form of neutral fat, fatty 
acids, or soaps. The more difficultly melting neutral fats are present 
usually as white or yellow lumps, scales or droplets, depending on their 
melting-point. Fatty acids appear usually as short, delicate, curved needles 
which occur in such thick masses that the shape of the individual crystal 
can seldom be made out. The soaps, on the other hand, crystallize out in 
long needles which are arranged in clusters or fans, or in short plump 
crystals, or scales. That the most of the needles seen in a stool are soaps 
may be proven by examining the stool after it has been extracted with 



THE INTESTINAL CONTENTS AND THE FECES 391 

ether. The droplets of neutral fat are soluble in ether; the fatty acids are 
dissolved on warming and in ether, while the soaps are not dissolved on 
warming nor are they soluble in ether unless they are first split by acid. 
An easy test for neutral fat is to mix the specimen under the microscope 
with i drop of a concentrated alcoholic solution of Sudan III which has 
been filtered just before using. The droplets take a color which varies 
from orange to a blood-red color while the soaps and the fatty acid crystals 
remain unstained. 






Acholic stools usually contain much 
fat in crystals which are mixed homo- 
geneously with the fecal matter. Such 
stools have a glistening gray appearance 
and microscopically contain large num- 
bers of fat droplets and large masses of 
fatty acid crystals. 

In diarrhea the masses of fat needles 
are sometimes large enough to be seen 
with the naked eye as minute points. 
Sometimes, as in pancreas disease, the 
lumps of fat in a stool are even the size of 
a nut and of a whitish-gray or a yellowish 
color like tallow; or the fat may be evacu- 
ated as a melted oil which hardens over 
the cold stool. Indeed, the whole stool 

may resemble oil. In a recent case of ', ' 

probable cancer of the pancreas the stool .' j 

looked like a mass of vaseline. ■ . \ \\ 

These excessively fatty stools are met / . UM\ 

with when the diet contains an over- 
supply of fat. The stools of patients 
on the olive oil treatment for gall-stones 
may contain firm lumps of saponified l 

. . • Fig. 73- — A sheaf of huge fatty acid crystals 

tat Which Vary 111 Size from that OI a seen often in stools after they have stood a 

pea to that of a hazelnut. Small firm 

masses of fat are found in the stools after a meal containing fats with a 

high melting point, as pork, mutton, or tallow. 

Pathologically the stools are fatty when, because of disease of the 
intestinal mucosa or of the lacteals, the fat cannot be absorbed, as in atrophy 
of the mucosa, in amyloid disease and in cases with extensive tuberculosis 
of the retroperitoneal lymph-glands. This last mentioned disease (tabes 
mesenterica) is the most common cause of very fatty stools in cases without 
jaundice. In fact, in many doubtful abdominal cases the inspection of the 
stools alone would suggest this diagnosis. The stool may be fatty in peri- 
tonitis and even in simple catarrh preventing absorption. There is a fat 



392 CLINICAL DIAGNOSIS 

diarrhea due to various diseases of the small intestine which is often con- 
fused with " diarrhea pancreatica." 

In cases of obstructive jaundice from 55 to 78% of the fat ingested will 
be lost in stools (normally from 6 to 10%) . Acholic stools in cases without 
jaundice may contain large amounts of fat. The cause of this condition 
is in doubt. Some believe that bile secretion may be temporarily sus- 
pended, others that bile pigment is present but has been changed to some 
colorless forms. 

In pancreatic disease fatty stools are common, but to be of value in 
diagnosis they must be exceedingly fatty. It is, however, also true that in 
severe pancreatic disease they may not be fatty at all provided the fat 
ingested had already been emulsified. Muller showed that while normally 
84% of the fat is split in the bowel yet if the pancreatic juice be excluded 
only about 40% will be split. Others consider that 80% may be split 
but not saponified. The diagnosis of pancreatic disease is exceedingly 
difficult and cannot be made from the fatty stool alone, yet if all the 
elements of LeNoble's symptom complex are present (no jaundice, gly- 
cosuria, much fat in stools, many fatty acid crystals but no soaps, no 
hydrogen sulphide, skatol or indol, the stools rancid but not putrid and 
with few bacteria) one is very safe in assuming the existence of some 
pancreatic trouble. 

Estimation of-Fats and Soaps. — The stool is first evaporated over the water-bath 
until of a semisolid consistency, then mixed with about 50 c.c. of absolute alcohol, again 
evaporated and this repeated until it is perfectly dry. A certain amount is then powdered, 
dried at ioo° C. and weighed. This is rubbed up with sand and extracted for from 8 
to 10 hours with ether. The ether residue is washed with warm water, dried in a desic- 
cator and weighed. It consists of neutral fats and fatty acids. 

To isolate for weighing the neutral fat the residue is again dissolved in ether and 
shaken out with a dilute soda solution, which will remove the fatty acids. 

The fatty acids are determined by dissolving a weighed amount of the ether residue 
in alcohol and ether and then titrating this with an alcoholic solution of potassium 
hydroxide, phenolphthalein used as indicator. 

For the determination of the soaps, some of the ether residue is boiled with acid 
alcohol, dried, extracted again with ether and the free fatty acid in this extract titrated 
as above. 

If one wishes to determine at once the neutral fats, the split fats and the soaps, 
the stool is first boiled with acid alcohol before it is extracted with ether (Muller). 

Mucus. — The stools always contain a certain amount of mucus (Boas). 
This is, however, seldom seen even microscopically and must be tested 
for chemically. Any visible mucus is somewhat abnormal. The mucus 
present is pure mucin, hence can be clouded by acetic acid. A specimen 
of stool is rubbed up with water, an equal amount of lime water added, it 
is allowed to stand for several hours and then acetic acid is added. A cloud 
will indicate mucus. 

Mucus is increased physiologically by hypersecretion, and forms a 
glassy or cloudy coating over hard fecal masses evidently to protect the 



THE INTESTINAL CONTENTS AND THE FECES 393 

mucous membrane against these. It may also be present after an active 
purge. This mucus is poor in cells. 

Mucus from the small intestine is intimately mixed with the stool and 
hard to isolate. In diarrhea these small flecks can be picked out with a' 
needle and if the stool be solid they appear as shreds or lumps which never 
are bile-stained. Such small flakes resemble those from the stomach. They 
are cloudy since rich in the detritus of digestion and in cells the bodies of 
which often are well digested or represented by masses of bilirubin granules 
and crystals. The so-called " sago granules," or " spawn -like " masses 
of mucus described by Virchow, are, Boas thinks, very rare. Mucus may 
be seen microscopically in the stool as small transparent lines and masses. 
The minute yellowish or greenish mucous granules or " islands " empha- 
sized by Nothnagel as indicating catarrh of the small intestine are, Boas 
and Schmidt consider, exceedingly rare and consist more of albumin than 
of mucus. Much mucus is present in the stools in cancer of the rectum with, 
stenosis (see also page 423). 

Some stools consist chiefly or entirely of mucus, which is glistening and 
jelly-like and is evacuated sometimes in masses resembling " frogs' eggs," 
or in strips sometimes over a foot long suggesting to the uninitiated tape- 
worms or pieces of bowel. This mucus comes from the large bowel, especi- 
ally the sigmoid flexure. Its evacuation is usually preceded by a colic 
which often is very severe and which leads to serious error in diagnosis. 
Some writers have attempted, but in vain, to distinguish between enteritis 
membranacea (a mildly inflammatory condition) and mucous colitis (a secre- 
tory neurosis). Excluding the cases with a pelvic tumor pressing against 
the rectum this condition is in large measure a secretory neurosis although 
the most of these patients have been constipated for years. Over 80% of 
these patients are women. Some pass mucous stools daily for a week at 
a time, some pass 1 a week or 1 a month, others pass them still more seldom. 
The relation between these stools and intestinal sand is interesting (see 
page 400). In general the student should be warned against advising 
operation on any patient who passes mucus in the stools if pain is the chief 
indication for the operation. 

Blood. — It is of course necessary to exclude the blood from raw meat 
and that due to hemorrhage from the mouth, nose, lungs and vagina. 
The presence of blood may be suspected from the red or tarry black color 
of the stool (although even 5% of blood in the stool may pass unnoticed) 
or it may be found microscopically or proven by chemical tests. Its dis- 
tribution in the stools is important; fresh blood covering a formed stool 
indicates hemorrhoids ; if evenly distributed with the food matter, it indi- 
cates hemorrhage into the stomach or upper bowel, providing the stool is 
solid. Tarry blood is usually from the stomach and duodenum; blood 
from small intestine (as in typhoid fever) is usually red. Bloody mucus 
suggests dysentery. Stools of blood and mucus without fecal matter sug- 



394 CLINICAL DIAGNOSIS 

gest intussusception and volvulus. Traces of blood are continuously pres- 
ent in the stools of patients with malignant diseases located anywhere 
along the alimentary tract ; traces present during definitely limited, usually 
short, periods may come from peptic ulcers, seldom from tuberculous ulcers. 
In typhoid fever blood may be detected chemically in the movement which 
precedes the bloody stool. In general chronic passive congestion there 
usually is blood in the stools, but not often in cases of xirrhosis of the 
liver with portal obstruction. It is present also in some cases of gastric 
hyperacidity and usually in poisoning by mercury. 

The tests for occult bleeding have their greatest value in the 
diagnosis of malignant diseases of the alimentary tract and in the differ- 
ential diagnosis between peptic or duodenal ulcer and nervous gastralgia. 

Guaiac Test. — About 3 gms. of feces are thoroughly mixed with a little 
water until fluid, then about }{ volume of glacial acetic acid added and it is 
then extracted with 10 c.c. of ether. To avoid emulsifying the mixture 
the tube should be slowly inverted, not shaken. 

A piece of clear brown gum guaiac (any green portions should first be 
removed) about the size of a cherry is crushed, and dissolved in a test-tube 
half full of alcohol. This tincture should have a light cherry color. 

To about 5 c.c. of the extract of the stool are added about 0.5 c.c. of 
the guaiac tincture and then 1 c.c. of commercial (3%) hydrogen peroxide, 
or an equal amount of old oil of turpentine. Fatty stools should first be 
extracted with ether to remove the fat. In the presence of blood a blue 
color quickly develops which first deepens and then fades to a pale green. 

If the diet contains raw meat this test may be positive, but seldom, if 
ever, if the meat is cooked. Eggs never disturb the test. The fact that the 
acid stool has been extracted with ether eliminates the most of the disturb- 
ing factors, such as milk, pus, saliva, spices and all drugs containing iron. 

The Aloin Test of Klinge and Shaer is very delicate. As a preliminary 
step all foods containing hemoglobin and chlorophyll and all drugs should 
be discontinued and the patient put on a milk, bread, eggs and fruit diet. 
Much fat with the food should be avoided. The diet period is to be 
limited by charcoal, not by carmine. 

The stool if very dark in color is rubbed up with 10 volumes of alcohol, 
this filtered off to remove the urobilin and the stool dried on the filter paper. 
About 5 gms. are then digested for 1 or 2 minutes with 5 c.c. of ether. From 
1 to 1.5 c.c. of oxygenated turpentine are then superimposed and 0.5 c.c. 
of fresh 3% aloin solution (0.3 gm. powdered aloin is dissolved in 10 c.c. 
of 60 to 70% alcohol). A fine red ring appearing in from 3 to 5 minutes 
at the line of separation indicates blood if the patient has been on the 
above-mentioned diet for several days and if the test is confirmed by 
several examinations. 

Benzidin Test. — Method of Schlesinger and Hoist. A piece of feces about 
the size of a pea is thoroughly mixed in a test-tube about % full of water, 



THE INTESTINAL CONTENTS AND THE FECES 395 

using a clean glass rod, and the mixture brought to the boiling point over 
the free flame to destroy any enzyme present. While this is cooling a fresh 
approximately saturated solution of benzidin is made by dissolving ina clean 
test-tube a knife-point full of benzidin purissimum (Merck) in about 2 c.c. 
of glacial acetic acid. One now mixes in a clean test-tube 10 or 12 drops 
of this fresh benzidin solution and from 2% to 3 c.c. of commercial hydrogen 
peroxide (3% H 2 2 ), shaking the tube lightly. If a green or blue tint 
appears in this reagent it cannot be used. If the reagent is satisfactory, 
a few drops of the boiled suspension of feces are added. If blood is present 
a beautiful green, bluish green, or blue color will appear within 2 minutes 
which changes later to violet. The depth of the blue and the rapidity 
with which it appears will depend on the amount of blood present. Only 
a definitely green or blue color is positive; faint tints are to be disregarded. 

This test performed in this manner requires less than 5 minutes and is 
by far the most sensitive of all. When negative it excludes the presence of 
even minute traces of blood. If positive, it should be confirmed by the 
guaiac test, which is safer if meat has not been entirely excluded from the 
diet. In all tests for blood 2 or 3 portions of the same solid stool should be 
used, as one may contain considerable blood and another none. 

Vaughan 8 recommends for clinical use Wagner's dry benzidin test for 
blood which he has slightly modified. A knife-point of powdered benzidin 
(an amount the size of a match head) is mixed in a short tube with 2 c.c. 
of glacial acetic acid and 20 drops of a 3% solution of hydrogen peroxide. 
This reagent will remain good but for 2 or 3 hours. A particle of feces the 
size of a match head is picked up on a toothpick, spread somewhat on a 
glass slide and covered with 1 or 2 drops of the reagent. The presence of 
blood will be indicated ty the appearance in 5 seconds of a greenish blue 
color which persists for a minute or more and then fades. It is best seen 
if the slide is viewed against a white background. 

Instead of a slide a piece of glazed paper (a calling card) may be used 
and has the advantage that it need not be cleaned up afterwards. 

This test is not, says Vaughan, too delicate for clinical use and is not 
interfered with by meat fibers, pus and the usual drugs and foods, but 
would be positive if the diet contains considerable raw meat. 

Pus. — Very rarely is there enough unaltered pus in the stools to be 
recognized macroscopically and when there is it always indicates the 
rupture of an abscess (e.g., appendix abscess) into the intestine. On the 
other hand the contents of even large abscesses may be passed unrecognized, 
so altered may the pus be by digestion and decomposition. Pus-cells are 
not recognizable microscopically if mixed with food, but are when enclosed 
in masses of mucus. The few scattered pus-cells seen in most mucus have 
no significance (although the mucus may have) , since the normal intestinal 
mucosa contains many leucocytes which wander into the lumen of the 

8 Jour, of Lab. and Clin. Med., March, 1917, vol. ii, No. 6. 



396 CLINICAL DIAGNOSIS 

bowel. Mucus containing an unusual number of single pus-cells may mean 
catarrh; that containing masses of these cells means ulcer or (especially 
if it contains blood also) cancer. 

Muscle and Albumin. — Muscle fibers can be found in the stools of all 
normal persons on a meat diet. The degree of their digestion may be esti- 
mated by their appearance. Some show beautiful cross, others only a 
longitudinal striation, while still others can be recognized as muscle fibers 
only by their shape, size, and yellow color due to their affinity for bilirubin. 
Under normal conditions these fibers occur singly. To find bundles of 
muscle fibers means a pathological increase. In diabetes and diarrhea 
they appear in the stool in large numbers, even in masses visible to the 
naked eye. The question whether or not there is a pathological increase 
of the muscle fibers in a solid stool is best answered by noting their size, 
shape and striation. In general it may be said that under normal conditions 
one finds none in bundles and none with the cross striation well preserved. 

The condition of lientery (the presence in the stools of grossly visible 
particles of undigested food) is, of course, present in cases with a gastro- 
intestinal anastomosis, but also in a great variety of other conditions with 
loose bowel movements. 

The presence of an abnormal amount of muscle-fiber in a fairly thick 
or solid stool and without diarrhea is known as azotorrhea. This is suggestive 
of, but not at all conclusive of, pancreatic disease. 

Milk curds and masses of coagulated albumin may be found in the 
stools of infants, but also of adults on a pure milk diet or one containing 
much coagulated egg. Of the so-called milk curds found in infants' stools 
the tougher ones consist of casein, while many of the softer ones are almost 
pure fat. 9 

Starch. — It is seldom that well-preserved single starch granules are seen 
in the stool of an adult, yet vegetable masses enclosing masses of such 
granules are common enough. The presence of the former indicates either 
diarrhea or hyperacidity. Starch granules are never bile-stained. In 
achylia pancreatica the starch may not be increased in the stools since the 
bacteria will break it up. The lack of bile also may cause no increase. 
The iodine test will indicate the extent to which starch granules have 
been digested, a blue color indicating the unchanged granules; red, a 
slight digestion. 

Carbohydrates. — To detect carbohydrates in the stool Strauss recom- 
mends the following method: From 2 to 3 gms. of the dried stool, which 
should not contain mucus or lactose, are heated for an hour and a half in 
a flask with a return cooler, with 100 c.c. of 2% HC1. The contents of the 
flask are then cooled, neutralized quite accurately with sodium hydroxide, 
filtered through an asbestos filter, washed with water, if necessary filtered 
a second time and the filtrate brought up to 200 c.c. Fifty cubic centimeters 

9 Jour. A. M. A., 1910, p. 372, and Talbot, Arch, of Pediatrics, Dec, 1909. 



THE INTESTINAL CONTENTS AND THE FECES 397 

of this are poured into a 300 c.c. beaker and the sugar determined quanti- 
tatively. This amount multiplied by 0.94 equals the amount of starch 
originally present. To test the stool qualitatively for glucose it may be 
boiled with water and the filtrate then tested with Fehling's or similar solu- 
tions. The albumin should first be precipitated with the acetate of lead, 
the lead then removed with C0 2 and the nitrate tested. It is seldom that 
any glucose is found in a stool which has not been boiled with acid. 

Ferments. — The ferments in a stool may be extracted with glycerin 
and the digestive power of the extract tested. Leo's method is to mix 
the feces with chloroform water until they form a thin pasty mass and to 
suspend in this a gauze bag. containing from 2 to 5 gms. of finely divided, 
previously boiled blood fibrin which will absorb the ferments. In 24 hours 
the bag is removed, the fibrin washed a number of times with water and 
then tested for the ferments. To test for trypsin, a little of the fibrin is 
placed in a 1% solution of soda and left in an incubator for a few hours 
and then filtered. A positive biuret will indicate the presence of trypsin. 
For diastase a little of the fibrin is placed in a thick starch solution in the 
thermostat and in a few hours its filtrate tested with dilute Lugol's. If the 
first drops do not give the blue color of starch then some has been digested. 
Normally the ferments are destroyed or absorbed in the intestine, yet 
each may be present in the stools in cases of diarrhea. 

Microscopy of Stools, — For the microscopical examination of the stools 
care must be taken in the selection of the particles to be examined, for a 
random search usually yields little. To examine for parasite eggs, etc., 
it is best to mix the stool with water and centrifugalize it or allow it to 
sediment in a tall jar since these will sediment rapidly. Mucous particles 
from the fresh, still warm stool should be chosen if protozoans are the object 
of search. In searching for blood especially it often makes a considerable 
difference whether the right particle is chosen or not. 

Epithelial Cells. — Squamous epithelial cells are often found in the 
mucus on the surface of the stool, and come from the anal regions. Great 
numbers may be present in cases of rectal cancer and of proctitis. 

Cylindrical epithelium is the commonest form of epithelial cell found. 
In some cases of diarrhea they are so numerous that these cases are grouped 
under the title " desquamative catarrh." The cells are easiest found in 
the mucus obtained by lavage of the rectum and sigmoid. They show all 
grades of degeneration. Some, even the goblet-cells, are fairly well-pre- 
served, others are very fatty, while still others are merely remnants of cells. 

Crystals. — Triple phosphate crystals, irregularly formed as a rule, and 
typical calcium phosphate crystals are usually present. In addition are 
found calcium salts of still unknown acids which take the form of irregular, 
oval, or circular masses, sometimes fissured, sometimes concentrically 
striated. These are always bile-stained. The masses of calcium soaps are 
frequently numerous (see Fig. 65). 



398 CLINICAL DIAGNOSIS 

Cholesterol is often present in the stool, but rarely in typical form and 
must be demonstrated by chemical methods. Char cot-Ley den crystals 
(Fig. 74) are met with in a great variety of diseases. If present, in abun- 
dance they always suggest the presence of some animal parasite, it may be 
any from the harmless oxyuris to the pernicious uncinaria. See Fig. 87. 
Bismuthous suboxide occurs as black, irregular rhombic crystals after 
the administration of bismuth subnitrate (see Fig. 76) . Hematoidin crystals 
are met with but are very rare. 

Remnants of undigested food make up the most of the microscopic 
picture. The student's attention is especially attracted by the thorn-like 
spines (see Fig. jj) from various fruits and berries; the spiral cells from 
the veins of leaves; cells with thick cellulose shells, some resembling soap 

masses, others very like parasite eggs ; 
the elastic tissue from meats, etc. The 
list is too long and varied to allow 
enumeration (see Figs. 75, 76). 

Macroscopical Examination — Gall- 
stones. — To find gall-stones in the 
stools (and a careful search should be 
continued for 15 days after an attack 
of colic) the stools are well mixed with 
water and then rubbed through a sieve. 
The failure to find a stone after a quite 
typical attack of colic may, granting 
fig. 74.— Charcot-Leyden crystals from the the pain was due to a gall-stone, have 

stools. X 400. r fe ' 

several explanations : the stone may not 
have entered the cystic duct, but fallen back into the gall-bladder; it may 
have remained in the ampulla of Vater; or, it may have disintegrated 
in the bowel, as some, perhaps all soft stones without a hard rind, do. 

Gall-stones vary in size from tiny concretions to others as large as hen's 
eggs. The single stones are usually spherical and have a rough surface, 
but when multiple have smooth deep facets. When fractured they may 
be seen to consist of concentric layers. (Every suspected mass in the stool 
should be fractured since enteroliths and fragments of bone, as, e.g., a 
bird's vertebrae, sometimes closely resemble gall-stones.) Gall-stones are 
composed chiefly of cholesterol and the calcium salt of bilirubin (sometimes 
also of biliverdin, bilihumin, bilicyanin), together with traces of calcium 
carbonate. Some rare stones consist of almost pure cholesterol. These are 
glistening gray in color, rough and soft. 

For analysis the stone is dried and powdered. This is necessary even 
in the case of small stones since they have a mucous coating which prevents 
the action of the solvents. This powder is then dissolved either in alcohol 
and ether, in which case the cholestrol crystallizes out as the ether evapor- 
ates, or in boiling alcohol from which the cholesterol will precipitate on 




f 



-■ -^ ; , / 



h. 



^- 






If' 






\ 



\ 



/ 



\ 



k 



4 



/V 

I \ 



\ 



Fig. 75. — a, Vegetable cells in stools, resembling parasite eggs. The cell on 

the left is an unfertilized ascaris egg; b, lycopodium spores; the crystals are 

an iron salt. X 400. 



; V 

.v - 


x - _ < 


V 


r 






; > 1 

1 ^ 


t T* *#^* 


\ 


- 1 


I B ' " ' ' 


s * 


1 





Fig. 76. — Cells in stools. A, B, muscle fibers; C, D, vegetable cells; E, F, spinal fibers from 
a piece of lettuce; G, cellulose framework of vegetable tissue. The crystals are of bismuthous 

oxide. X 400. 



THE INTESTINAL CONTENTS AND THE FECES 399 

cooling. After the cholesterol is extracted the residue is treated in the cold 
with very dilute KOH solution which will extract the bilirubin in a yellow 
solution which will give Gmelin's test. This solution will be blue if bili- 
humin is present. 

Pseudo Gall-stones. — A little care would prevent the many mistakes 
which result from a failure to distinguish between true and pseudo gall- 
stones. Each suspected concretion should at least be fractured and many 
tested chemically. The more dangerous pseudo gall-stones are masses of 
vegetable tissue, seeds of fruits, pieces of bone, enteroliths, and masses of 
fats and soaps of high melting point. Olive oil won its great reputation as 
a medicine to remove gall- 
stones since if administered in I 
large doses the stools may con- 
tain even hundreds of firm 
masses of soaps. I - 

Gall-sand. — The sand-like ^T 

sediment often called gall-sand / ' 
and so abundant in f some stools, . 41 
probably does not come from | 
the gall-bladder. Genuine gall- 
sand would probably disappear 

in the bowel, and even though • - 

it did not the bile could not ex- ■ ■ 

plain the large quantities found 
in some stools (Naunyn). And „ c . " - . IT""... „ A < L A1 _ . ,[ 

y J ■ .' riG. 77. — Spines forming the down, that on the right 

Vet gall-Sand may be recovered of a raspberry, on the left of a quince. These are often 
J & J taken tor the embryos of parasites. 

from the duodenum by Lyon's 

method (see page 385) and it certainly explains some cases of parox- 
ysmal hyperchlorhydria. 

A private patient, a physician 45 years old, had, for over 20 years, suffered quite fre- 
quently from "heart-burn, "often paroxysmal and particularly aptto waken him at 2 A.M., 
at which time he often would induce vomiting thus obtaining complete relief. He could 
always gain relief also by taking soda. The diagnosis made was gastric ulcer, although 
the rontgenological examination showed the emptying time of the stomach normal 
or even shortened. 

On one occasion he had pain, not very severe, suggesting a gall-stone in the cystic duct 
and suggested at once operation. The gall-bladder and its contents appeared at first 
normal but more careful examination revealed fine granules of gall-sand. No evidence 
of ulcer was found. The gall-bladder was drained. This operation was followed by 
great relief. 

Pancreatic stones are seldom if ever found in the stools. Those in 
the pancreatic duct are white, usually single and consist chiefly of calcium 
carbonate. 

Enteroliths. — By enterolith is meant a hard lump of food or mass of 
hardened feces encrusted with inorganic salts. Enteroliths are seldom 



400 CLINICAL DIAGNOSIS 

passed in the stools. Their chief importance is in connection with appen- 
dicitis, although the little hard lump of merely dried feces often found in 
the appendix seldom is an enterolith. 

Intestinal Sand. — Intestinal sand is the name applied to the sediments 
of small, stony hard concretions about the size of genuine sand grains which 
occasionally is found in the stools. Concretions over 2.5 mm. in diameter 
should not be called " sand." It sometimes appears in considerable quan- 
tities, even half an ounce in 1 stool. Its passage may be an incident of 
a nervous crisis arid may be preceded by considerable pain. Most of the 
cases reported have been neurasthenic patients with a history of mucous 
colitis. (This is not surprising, since we seldom examine the stools of 
normal persons.) 

Many specimens of so-called intestinal sand have proved to be pseudo- 
sand made up of the seeds of berries, bananas, granules from the seed case 
of pears (these vegetable masses can be easily recognized by studying the 
cross-section of a granule), concretions of altered blood-pigment, bile- 
pigment and concretions of medicine, as salol. In other cases the " sand " 
is genuine, i.e., is quartz swallowed with the food. The best article on this 
subject is that of Myer and Cook, 10 who believe that most of the sand is 
vegetable matter. They cite a case in which the granules which were of 
stony hardness proved to consist of resin and tannin, products, of the diges- 
tion of the milk-cells of bananas. But apart from these cases one does in 
rare instances meet with a condition which would seem to be a secretory 
neurosis which deserves the name " gravel-forming enteritis " (Eichorst). 

Chemical analysis of true intestinal sand has shown that it consists of 
the phosphates and carbonates especially of calcium, but also of magnesium, 
iron, etc.; while in some of the granules calcium sulphate predominates. 11 
Practically all, however, contain some organic matter, as bacteria, fat, 
cholesterol and urobilin. The granules are described as spherical or angu- 
lar in shape, very hard, from 0.15 to 2.5 mm. in diameter, and often of a 
reddish-brown or green color. 

We have seen several cases of pseudo-sand and 2 very good cases of, we believe, 
real intestinal sand. In 1, a young boy ill with an indefinite nervous disorder, such 
large amounts of fine granules were occasionally passed that they made up a very con- 
spicuous constituent of the stool. The other patient was a young woman with an intes- 
tinal neurosis. In this case the granules seemed to be plugs of cells impregnated with 
carbonates. The nature of these cells could not with certainty be determined, but they 
were the size of columnar epithelial cells. We have found a few such granules in simple 
diarrheal stools and the further study of such cases may determine the nature of these 
interesting bodies. 

Bedford 12 thinks that in his case there was a definite relationship 
between intestinal sand and gout and tophus formation. 

10 Am. Jour. Med. Sci., March, 1909. 

11 See also Garrod, Lancet, March 8, 1902, and Eichorst, Deut. Arch. f. kl. Med., 
1900, Bd. 68, page 1. 

12 Lancet, July 26, 1902. 



THE INTESTINAL CONTENTS AND THE FECES 401 

Tumor Fragments. — Tumor fragments and adenomatous polyps 
(which may develop as an independent disease or grow in the neighborhood 
of cancers or ulcers) may be met with in the stools as firm fragments of a 
grayish-red color and tough consistency. These have their origin in the 
rectum or colon, or even higher. They are very easily overlooked unless 
the stool is thin. They may be recognized from the general arrange- 
ment of the nuclei in the microscopic sections; the fine details all will 
have been lost. 

Intestinal Parasites. — Protozoa. — Rhizopoda. 

Entameba Dysenteries or Histolytica. — The protozoon formerly called 
Ameba coli (and still so called by many), is now generally admitted to be 
the cause of " amebic dysentery," a colitis characterized by its very chronic 
course, a tendency to relapse and the frequency with which it is associated 
with abscess of the liver. When ^^^^^^^^^^^^^^^^^^^^— 
Shiga's bacillus was first discovered 
it was thought by many to be the H 
cause of the so-called amebic dysen- (TV.. * 

tery and Ameba coli was considered 
a secondary invader; but amebic 
dysentery and bacillary dysentery 
now are recognized as two distinct mm 
diseases. ' M 

The ameba? are often abundant 
in the masses of bloody mucus 
passed in the stools of patients with „.-■.,,, 

Fig. 78. — Ameba coli (Entameba dysentenae) , 

amebic dysentery, although many common form, x 400. 

more are in the ulcers which undermine the mucosa of the colon and of the 
ileum and in the sinuous fistulas which radiate from these ulcers for long dis- 
tances under the mucous membrane. They are found also in the contents of 
but more easily in the walls of the liver abscesses which so often complicate 
this form of dysentery, and in the sputum of patients through whose lungs 
such abscesses have ruptured. 

Entameba dysenteric (Fig. 78) is a rhizopod which varies in diameter 
from 8 to 50JU. It has a clear hyaline ectosarc, seen best in the pseudopods, 
and a finely granular endosarc which usually contains some of the para- 
site's ingesta (red blood-cells, leucocytes, bacteria, epithelial cells and 
particles of food) and often 1 or more vacuoles which do not pulsate. Its 
spherical nucleus, about 6/jl in diameter, is sometimes, although rarely, 
seen in the living parasite. To demonstrate the nucleus one kills the 
organism with corrosive sublimate and stains it by appropriate methods. 

The organisms found in various cases of this form of dysentery do not 
all look just alike. Some differ so much in appearance from those in 
others that the temptation is ever present to describe different varieties 
of the parasites. 
26 



402 CLINICAL DIAGNOSIS 

In the fresh stool, if the stage of the microscope is not too cool, this 
parasite is ameboid. Some move slowly, others so fast that they are with 
difficulty kept in the field of vision, while others merely project i of their 
pseudopods . They are very sensitive to an acid reaction of their environment . 

These amebae multiply by simple division. Resting, resistant forms, or 
" encysted amebae," the nuclei of which have divided into several nuclei 
each in the center of a clump of protoplasm, have been described. It is 
probable that the infection of a new host is effected by these encysted forms. 

Since it is almost impossible when examining a stool to distinguish 
resting amebae from swollen degenerated epithelial cells it is of great im- 
portance that the diagnosis be based solely on cells which unmistakably 
project pseudopods and never on those the motility of which is doubtful 
no matter how closely they may resemble amebae (although quiet cells 
resembling amebae which contain red blood-cells, leucocytes and bacteria 
probably are amebae). 

The stools should be examined while fresh and while warm for the para- 
site is sensitive to cold. If the specimen is kept warm the parasite will 
remain active for even 24 hours. (The common mistake of overheating 
the stool should be avoided.) 

In choosing particles for examination one selects, if present, flecks 
of mucus and in the absence of these the liquid part of the stool. If the 
stools are firm the patient is given a dose of salts and the next liquid 
stool examined; or, a solid fragment of stool may be mixed with normal 
salt solution and then examined, One of the best methods of obtaining 
this parasite is to pass a rectal tube and examine the little fleck of mucus 
which the edge of the eye of the tube will scrape from the mucosa. The 
parasites if present at all will usually be found in the mucus in clusters 
of scores to a field. 

During an acute exacerbation of a case of chronic amebic dysentery the 
patient usually passes 5 or 6 stools a day, seldom more. The stools are then 
loose, not watery, and contain mucus which is usually blood-stained and 
generally mixed with some free blood. During the periods of constipation 
which separate these periods of diarrhea the amebae can often be found in 
the firm stools. Some cases of amebic colitis give no history of dysenter}^ 
but rather of years of constipation. This is particularly true of the cases 
complicated by amebic liver abscess. Other cases have throughout their 
entire course acute bowel symptoms, the frequent passage of small masses 
of blood-stained mucus. Still other cases are latent and cause the patient 
few, if any, symptoms, although the amebae in the stools may be numerous.. 

Years before Losch described Ameba coli as a pathogenic organism it 
was known that amebae were often present in the stools of persons who had 
no dysentery or ulcerative disease of the bowel; in simple diarrhea, in 
typhoid fever, in acute and chronic enteritis, colitis and proctitis, and even 
in the stools of healthy men. In 1893 Quincke and Roos described: Ameba 



THE INTESTINAL CONTENTS AND THE FECES 403 

coli (Losch), 15 to 2 s/jl in diameter (encysted forms 10 to 15/i), pathogenic 
to men and to cats; Ameba coli mitis, which is 25 to 35/x in diameter, which 
may ingest bacteria but never red blood-cells, and which is slightly patho- 
genic to man, causing a mild enteritis, but not at all pathogenic to cats, 
and Ameba intestini vulgaris, similar in appearance to Ameba coli but 
not pathogenic. 

The best contribution to this subject is that of Schaudinn 13 who sepa- 
rates Entameba coli from Entameba histolytica, the former the common 
harmless variety, the latter the pathogenic form causing dysentery. 

Craig claimed that Entameba coli can be found in the stools of 65% 
of normal persons after a dose of Epsom salts ; that it is somewhat smaller 
than Entameba histolytica (10 to 20^ in diameter), is less actively motile, 
has less difference between endosarc and ectosarc (the latter is less refrac- 
tile, the former has less demonstrable structure), vacuoles are less common 
and that the nucleus is more distinct than in the pathogenic variety. What 
is more important the pathogenic variety shows no encysted stage, but 
multiplies by sporulation. (For more details, see Craig. 14 ) 

The problem of non-pathogenic amebae may be of interest to the zoolo- 
gist, but the medical man should consider as possibly pathogenic every 
ameba he finds in the stools. Musgrave believes that any ameba which 
has been harmless may become pathogenic. Of the 300 persons in Manila 
whom he examined, 101 were infected with amebae. Of these, 61 had 
dysentery and the other 40 had no sign of the disease. During the next 
5 months, however, every 1 of these 40 developed a definite dysentery. 

Amebae may be cultivated, but it is with difficulty and only with certain 
bacteria. These cultures withstand drying for 15 months. 

Flagellata. — Of the flagellata only certain of the enflagellata are 
important in human pathology, especially the protomonadina and the 
polymastigina. Flagellated rhizopods and lower plants may be found but 
have no importance. 

Polymastigina. — These are flagellata with 3 equal or from 4 to 8 unequal 
flagella inserted at different points. Some have also an undulating mem- 
brane, often mistaken for a row of cilia. Of these 2 groups the Trichomonas 
and Lamblia are of importance. 

Trichomonas. — This is a. family of pear-shaped organisms, rounded in 
front, pointed behind, with at the anterior end 3 or 4 equally long flagella 
which often are united at their base. The undulating membrane, which 
is usually present but not always seen, begins at the anterior pole and 
extends obliquely backward. The nucleus is anterior and behind it are 
1 or more vacuoles which do not pulsate. A flagellate about to extrude its 
flagella may resemble an ameba since the protrusions of the cell membrane 
resemble pseudopods. 

13 Arbeit, a. d. Gesundheitsamte, 1903, xix, p. 563. 

14 Am. Med., May 27 and June 3, 1905. 



404 CLINICAL DIAGNOSIS 

Trichomonas Vaginalis (Donne). — This parasite (Fig. 79) is from 
15 to 25^ long and from 7 to 1.2/i broad. Its posterior end is drawn to 
a thread. Its cuticle is thin and its protoplasm free from granules. It 
has usually 3 flagella of equal length, which sometimes seem united at the 
base, and an undulating membrane the edge of which has probably been 
mistaken as the fourth which some describe. This parasite is found in 
abundance in the acid secretion of cases of vaginitis. 

In the intestine various similar flagellates have been described under 
such names as Protoryxomyces coprinarius, Monocercomonas hominis 
(Grassi), Cinaenomonas hominis (Grassi), Trichomonas hominis (Grassi), 
Cercomonas coli hominis (May) but all of these are now considered to be 
identical with Trichomonas vaginalis, which parasite can live in the urethra, 
the large and the small intestine, the stomach, the mouth, and even lung 

r— • ~ - — r ■ •- "*m cavities and in the Dietrich's 

_ .'-• " plugs. It owes its name to the 

fl K s Pf \ fact that it was discovered first 

-VI S / % : ; f -\ ; i n the vagina. It has long 

been a debated question 

: ' S| ^V -\ wh^her these parasites were 

'■•■■■} ' . .X ■■■■- . I harmless or not; whether they 

./.'-■. -'.'"'- . caused diarrhea or merely 

^- • - - — *• - ■ • •• -'■"— — ~^— ^3 aggravated a trouble already 

fig. 79.— Trichomonas vaginalis. present. It is now considered 

pathogenic and the adequate cause of even a severe diarrhea. 

J. W., aged 58, male, Med. No. 10101, was admitted Aug. 11, 1920, with a severe 
and persistent diarrhea of 3 years' duration. One year ago a physician discovered that 
his blood Wassermann was 4 plus and since that time he has had thorough anti-syphi- 
litic treatment with both neo-arsphenamine and mercury, but the diarrhea has been 
uninfluenced by the treatment and the stools now are as frequent as 10 to 20 a day. 
Examination of the stools revealed the Trichomonas intestinalis in large numbers, 
4650 per c.mm. in 1 specimen. (In this count Dr. Hahn considered only definitely 
motile organisms. Had he included those not moving the count would have been at 
least 10% higher.) There was no blood and little mucus in these stools. Examination 
of the mucosa of the colon for amebae was negative. Gastric analysis showed a complete 
absence of free hydrochloric acid, pepsin was present, but no blood or lactic acid. Yeast 
cells were numerous. The olive tip of the Einhorn tube did not pass through the pylorus 
in 3 hours. The stomach practically emptied itself in 1 hour of a test meal consisting 
of 1 slice of toast and 12 ounces of water. Fluoroscopy showed very active gastric 
peristalsis and signs of pylorospasm; great quantities of gas in the colon; a large, long, 
fixed appendix, slightly segmented. The blood count was: red cells, 5,156,000 per 
c.mm.; white cells, 9,900 of which 2.27% were eosinophiles ; hemoglobin, 110%. Color 
index, 1.06. Blood Wassermann, negative. Rhamy and Metts 15 reported a group of 
these cases from the same county of Indiana. 

Lamblia (Fig. 80). — This family of pear-shaped organisms is character- 
ized by a deep concavity on the inferior surface and by 4 pairs of flagella, 

15 Jour, of A. M. A., Apr. 15, 1916, lxvi, p. 1190. 



THE INTESTINAL CONTENTS AND THE FECES 405 

3 on the edges of the concavity and i at the posterior extremity. The 
parasite found in man has been named: Lamblia intestinalis, Cercomonas 
intestinalis (Lambl), Cercomonas coli (May), Trichomonas intestinalis 
(Leuckart) . 

Its protoplasm is hyaline or very finely granular and never contains 
solid inclusions, it has a very fine cell-membrane and a nucleus which is 
dumb-bell shaped, situated at the base of the concavity. It has 4 pairs 
of flagella of almost equal lengths (9 to 14^), 1 pair on each side of the con- 
cavity, 2 pairs at the projection on the inferior edge of the concavity and 
1 pair at the end. The motile forms vary from 10 to 2iju in length and 
from 5 to 12^1 in width. This parasite lives in the jejunum and duodenum, 
each fastened on the top - - — 

of a columnar cell which • . , 

it embraces with its con- ■••• / ' 

cavity. In some cases 
they are present in such 
numbers that they form 

a membrane covering the V 

mucosa. Those which ..■-■"". < 

reach the large intestine ..'■ 
become encysted, taking ^ 

the form of round or oval ■ ■ }< 

bodies from 10 to 14/A / 

long and 8 to 10/x wide . • , • . 

with a very distinct mem- :'•*. ' , - - 

Drane Surrounding the p IG> go. — Lamblia intestinalis, showing the motile form in differ 
Organism. The motile ent positions, and stages of its encysting. X 900. 

parasite is seldom seen in the stools unless the patient has a severe diarrhea 
in which case they are seen thrashing about rapidly and very aimlessly. 
The stools of a patient who has taken a large dose of Epsom salts should 
be examined as fresh as possible and on a warmed stage. The number 
in the stools may be enormous; even, it is estimated, 18,000,000 in 24 
hours. They may be recognized by their concavity and their dumb-bell- 
shaped nucleus. Their most important hosts are the mouse, rat, rabbit, 
dog, sheep, cat, etc. Men are evidently infected from water. They have 
been found principally in children. Their pathogenicity is uncertain yet 
they may aid in producing the symptoms of other diseases and they cer- 
tainly thrive best in patients with intestinal troubles. 

It is always interesting to watch this organism encyst itself. It first withdraws its 
tail flagella, then becomes more oval until the concavity finally disappears, the flagella 
for a while projecting from its edges. In some cases the markings of the encysted form, 
commonly taken to indicate the folds of the parasite, seemed to be the edges of this 
closed cavity. A membrane could in some be distinctly seen. 

The protomonadina, or forms which have 1 or 2 equal flagella or 1 prin- 
cipal flagellum and 1 or 2 smaller ones, are much smaller than the above 




406 CLINICAL DIAGNOSIS 

mentioned polymastigina. Two of the 3 forms occur in man : Cercomona- 
didas, which have 1 flagellum and no undulating membrane, and the Try- 
panosomidae which have 1 flagellum and an undulating membrane which 
reaches the whole length of the parasite. 

Cercomonas Hominis. — These small flagellates, sometimes met with 
in the stools, are usually from 10 to 12/x, but vary from 8 to i6/a, in length. 
They are pear-shaped with 1 long flagellum even twice the body-length at 
the anterior end. Their motion is very rapid. Their pathogenicity is doubted. 
They have been found in other parts of the body also, including the sputum. 
Infusoria. — The infusoria are bilaterally symmetrical protozoans which 
have a permanent shape, are ciliated, which contain contractile vacuoles 
and usually a macro- and micro-nucleus. The order which is of most im- 
portance to us now is that of Heterotricha, which are uniformly ciliated, 
but with a border of longer cilia around the peristome. Of these the most 

important is the Balantidium group. 

Balantidium Coli or Paramecium Coli. 
— Balantidium coli is an oval parasite (Fig. 
1) from 60 to iooju long and from 50 to 70^ 
broad uniformly covered with cilia. The 
mouth, at the anterior end, is a funnel or cleft - 
Fig. 81.— Balantidium coii. shaped entrance which extends % the body 

(Copied from Braun.) ^^ ^ ^^^ fe smTOunded by ciHa 

about twice as long as those over the rest of the body. The ectosarc 
and the endosarc are clearly differentiated. The latter is finely granular 
and contains many fat or mucous droplets, starch granules, even red 
blood-corpuscles, leucocytes and bacteria. The nucleus is kidney- or bean- 
shaped and is accompanied by 1 or more accessory nuclei. Usually there 
are 2 contractile vacuoles which pulsate feebly. The surface is trav- 
ersed by parallel longitudinal lines connecting the 2 poles and most distinct 
at the anterior end. The anal orifice is at its posterior end which is rather 
blunter than the anterior. This parasite lives especially in the colon but 
in severe cases may be found even in the jejunum. It may be present in 
the stools in such enormous numbers that 1 drop of blood-stained mucus 
may contain 200 organisms. The pathogenicity of these parasites has 
been doubted but it is now agreed that they may be the cause of a very 
severe and stubborn catarrh which may even be fatal. (Some regard them 
as secondary invaders in cases of intestinal catarrh, but others on the con- 
trary believe that they can cause a catarrh which continues after they die 
out [Henschen].) Between 80 and 90 cases of apparently primary infection 
by these parasites are now on record, the most of them from Russia. A 
very good description of the condition is given by Strong and Musgrave. 16 
Klimenko l7 believes that they first cause a diarrhea by mechanically 

16 Johns Hopkins Hosp. Bull., February, 1901. 

17 Beitr. z. path. Anat. u. allg. Path., 1903, "Bd. 33, p. 281. 




THE INTESTINAL CONTENTS AND THE FECES 407 

irritating the rectal mucosa and later a catarrhal or even ulcerative colitis ; 
that they invade the intestinal wall, enter the blood-vessels and sometimes 
cause emboli to distant organs, but that their action is chiefly mechanical 
is indicated by the absence of any degenerative or inflammatory changes 
which would point to the action of a toxin. 

Enthelmintha — Trichina Spiralis. — The adults of trichina spiralis 
have not as yet been found in the stools of man. They may be studied in 
the intestines of rats, pigs, dogs, and cats. The male worm is from 1.4 to 
1.6 mm. long and 0.04 mm. wide; the female, 3 to 4 mm. long and 0.06 mm. 
wide. After the ingestion of infected meat the capsule of the encysted 
embryos is digested by the gastric 
juice and the liberated embryos ma- 
ture rapidly in the small bowel. On 
the second day the males die and the 
females bore their way through the 
mucosa either of the villi or at the 
base of the Lieberkuhn glands and lie 
in the lymph spaces where they vivip- (^\ 
arously hatch their young (which 

are O.O9 tO O.I mm. long, and 6/JL Wide) Fig. 82.— Ameba coli (Entameba dysenteriae). 

.. ., .. 111 -1 An uncommon, very hvaline, and verv ameboid 

mtO the lymph and blood-Stream. form of parasites, usually filled with red blood- 

/ _ 1 .. 1 1111 cells. The small forms are true ameba from a 

l hese travel paSSlVely in the DlOOd normal case. Entameba coli, drawn to the same 
.. .. 1 . .. scale. X 400. 

stream, then bore their way into the 

tissues and in 9 or 10 days come to rest in the muscles. They are then 
about 1 mm. long. A capsule forms around them, which in about 1 year 
begins to calcify (see Fig. 83). 

Ascaris Lumbricoides. — Ascaris lumbricoides, the ordinary " round 
worm," is so common an intestinal parasite, that it is found in the stools 
of about 0.4% of all persons examined (Garrison, Ransom, and Stevenson). 
The female is from 20 to 40 cm. long, 5 mm. thick, and has a straight and 
conical tail. The male is from 15 to 25 cm. long, 3 mm. thick, and has a 
posterior end which is bent ventrally into a hook and which terminates in 
2 spicules. The mouth of both sexes is surrounded by 3 papillae. The 
color of these worms is gray or a dirty reddish -brown. While this worm 
lives as a rule in the small intestine, so is usually met with in the stools, 
yet, especially in cases with high fever as typhoid, it may become very 
actively migratory, make its way into the stomach and appear in the 
vomitus, or crawl up the esophagus and appear in the nose or enter the 
Eustachian tube or crawl down into the trachea. To obtain the worm 
itself a dose of santonin will have a therapeutic as well as a diagnostic value. 
Its fertilized eggs (Fig. 84, d, e), which often appear in the stools in large 
numbers, are elliptical, from 50 to 70^ long and 40 to 50/x wide. (Those 
which we have measured varied from 65 to 80 by 45 to 55/x) and have an 
unsegmented protoplasm surrounded by a thick transparent shell, which 



408 



CLINICAL DIAGNOSIS 



v 



\VX 



x^ 



in turn is covered by a thick, gelatinous very lumpy envelope which usually 
is bile -stained. 

The unfertilized ova appear so different from the fertilized eggs that 
for a long time they were not recognized as eggs at all. Dr. O. T. Logan, 18 
who was one of the first to interpret them correctly, convinced us that the 
cell represented at the left hand edge of Figure 75 as a vegetable cell 

was certainly a typical 
unfertilized ascaris egg. 
/y ^^^ Houghton (personal com- 

munication) in a series 
of fecal examinations of 
500 patients in Wuhu, 
China, found that 71.2% 
were infected with asca- 
ris. Of these 38.6% 
passed fertilized and un- 
fertilized eggs, and 3% 
unfertilized eggs only. 

That the young worm 
from the recently hatched 
egg migrates through the 
intestinal wall and that 
some of these reach the 
lung and thence the bow- 
el via the trachea and 
esophagus thus infecting 
the host has been de- 
monstrated by Ransom. 19 ^ 
Oxyuris Vermicula- 
ris. — This little parasite 
(Fig. 86) occurs in the 
rectum and colon as high 
as the cecum where it 
inhabits the appendix. 
It may, however, travel 
even to the stomach and through the uterus and tube to Douglas's cul-de- 
sac. According to some it can bore its way through the intestinal wall 
and thus cause an abscess. About 0.8% of all adults examined are 
infected by this worm. These worms are white in color. The adult 
male is from 3 to 5 mm. long. Its posterior end is bent into a ventral 
hook. The female is 10 mm. long and 0.06 mm. wide. The eggs which 
are 50^ long and 16 to 20^ wide have a characteristic asymmetry. The 






Fig. 83. — Trichina spiralis, a, adult female. b, adult male. 
X 90. c, embryo. X 400. (I am indebted to Dr. C. L. Over- 
lander, of Boston, for these photographs.) 



18 Rep. Am. Soc. Trop. Med., 1908. 

19 Jour. A. M. A., Oct. 18, 1919, vol. 7s, p. 1210. 




,/ 



d 






Pig. 84. — Parasite eggs in stools, a, b, c, eggs of trichocephalus dispar, showing the different colors 
(species?); d, e, ascaris lumbricoides; d, envelope lost; e, perfect. X 400. 








FlG. 85. — Eggs of Tyroglyphus siro (cheese- or flour-mite), a, an egg magnified X 400 to allow a 
comparison of size with the eggs of Fig. 84. b, in the center an egg and above and below two mites 
soon after they hatched and had developed somewhat. X 100. Note. — We picture these eggs merely 
as a warning to the student that not all the eggs he may find in the stools are eggs of important parasites. 
One not infrequently finds eggs of the great variety of harmless insects, etc., which are swallowed with 
the food. When in doubt concerning an egg it should be carefully measured and then the attempt made 
to hatch it. If still in doubt the specimen should be sent to the Washington Laboratories (Department 

of Health). 



THE INTESTINAL CONTENTS AND THE FECES 



409 



parasite leaves the rectum at night to lay its eggs on the skin surrounding 
the anus and it is then that the itching occurs. The eggs when deposited 
contain a well-developed embryo. The skin around the anus should be 
scraped both for the adults and eggs, for it is rare to find them in the stools, 
except in the mucus which the stool gains on passing through the anal canal. 




Fig. 86. — Oxyuris vermicularis. A, B, and C, adults; A and C are females 
full of eggs. X 12. D, egg, X 400. 

Ankylostomum Duodenale and Uncinaria Americana. — These 2 
varieties of hookworm (Figs. 87-93) which belong to the nematode family 
Strongyloidse cause some of our severest anemias. They live in the duode- 
num, jejunum and ileum, sometimes thousands in 1 person but seldom 
more than a few hundred. They do not multiply in the bowel but the 



410 



CLINICAL DIAGNOSIS 



individual worms may live there for 5 years. They have recently attained 
great importance in this country through the demonstration by Stiles that 
they are the common cause of the " anemia of the South." In 500 cases 

chosen at random they were present in 3% 
of the individuals. 20 Stiles showed that there 
are in reality 2 varieties of hookworm in the 
Southern States, the form described years ago 
in Europe and another form to which he gave 
the name Uncinaria Americana. They have 
long been known as the cause of bricklayer's 
anemia, tunnel- workers' anemia, Egyptian chlo- 
rosis, miners' anemia, etc. The best descrip- 
tion of these parasites is that given by Stiles. 21 
Anchylostoma Duobenale, Uncinaria 

DUODENALIS, ANKYLOSTOMUM DuODENALE, 

The European Hookworm. — The body of the 
European hookworm is cylindrical (Fig. 87), 
its buccal cavity (Fig. 91) has 2 pairs of 
ventral teeth curved like a hook and 1 pair 
of dorsal teeth directed forward; the dorsal 
rib does not project into the cavity. The male 
is from 8 to 11 mm. long. It has a caudal 
bursa (Fig. 88) with dorso-median lobe and 
promiment lateral lobes united by a ventral 
The dorsal ray divides at a point % its 
length from the base, each branch being tridi- 
gitate. The spicules are long and slender. 
The female is from 10 to 18 mm. long. The 
vulva is at or near the posterior third of the 
body. The eggs (Fig. 89) are ellipsoid, 52 by 
33^, and segmented when 
laid. Development is direct 
without intermediate host. 
Necator Americanus, 
Uncinaria Americana 
(Stiles, 1902). — The Ameri- 
can form of hookworm dif- 
fers from the European in 
that its buccal cavity (Fig. 90) has a dorsal pair of prominent semilunar 
plates or lips and a ventral pair of slightly developed lips of the same 
nature, but no hook-like teeth. The dorsal conical median tooth projects 
prominently into the buccal cavity. The male is from 6 to 9 mm. long. Its 

20 See also Smith, Am. Jour. Med. Sci., 1903, vol. cxxvi. 

21 Eighteenth Annual Report of the Bureau of Animal Industry, 1901. 




I7. — Ankylostomum duodenale, natural size to right, much 
magnified male on left. (From Braun.) 



THE INTESTINAL CONTENTS AND THE FECES 



411 



caudal bursa (Fig. 92) has a short dorso-median lobe, which often appears 
as if divided into 2 lobes and prominent lateral lobes united laterally by 
an indistinct ventral lobe. The common base of the dorsal and dorso- 
lateral rays is very short. The dorsal ray is divided to its base, its 2 
branches being prominetly divergent and their tips bipartite. The spicules 
are long and slender. 

The female is 9 to 11 ^^-j^hipartiie 

mm. long, the vulva in /<Kdorsa/ /£ X t! P 

the anterior half of the 
body but near the equa- 
tor. The eggs are ellip- 
soid, 64 to 72^ long by 
36 to 40^ broad, some 
are segmented in utero 
and 'others contain a 
fully developed embryo 
when laid (Fig. 93). 

The hookworm's 
eggs (Fig. 89) found in the stools may be unsegmented but the majority 
have already divided into 4, 8, 16, etc. segments. They have a thin, clear 
shell. The older the feces and the warmer the weather the more ad- 
vanced will their segmentation be. This is especially true of Uncinaria 
Americana. 

To find uncinaria eggs it may be sufficient to mix a small particle of 




Fig. 88. — Caudal bursa of Uncinaria americana. (Schematic.) 



I 





<0&- 



Fig. 89. — Eggs of Uncinaria duodenalis. a, unsegmented; b, with four segments and showing 
nuclear spindles; c and d, later stages of segmentation. X 400. 

stool with a drop of water and spread it on a slide, but if they are not numer- 
ous it is better to follow the various suggestions of Pepper. 22 That is, to 
dilute the stools with about 10 volumes of water, strain it through 2 or 3 
layers of gauze in a funnel, and then centrifugalize it until the sediment is 
just thrown down. The supernatant fluid is now poured off, more water 
added, the tube is well shaken, the stool is again centrifugalized and this 
sediment examined. Again, use is made of the tendency of uncinaria 
eggs to stick to glass. A drop of the sediment is put on a glass slide and 
22 The Jour, of Med. Research, March 1908, vol. xviii, No. 1, p. 75. 



412 



CLINICAL DIAGNOSIS 



the slide is gently immersed in water. This will wash off much of the sedi- 
ment while the uncinaria eggs will stick. Another drop of the sediment is 
then added to the same spot and the slide again immersed. The process 
is repeated several times. In this way one may obtain fields which abound 





-Head of Uncinaria americana. 
(Schematic.) 



Fig. oi.- 



-Head of Uncinaria duodenalis. 
(Schematic.) 



in eggs. One disadvantage of this method is that the eggs of other parasites 

will be lost. 

The adults may be found in the sedimented stool following a small 

dose of thymol and then of oil. They are usually red from the blood which 

fills their intestine. 

Trichocephalus Trichiuris — Trichocephalus Dispar — Tricho- 

cephalus Trichuria — Whipworm. — The ordinary whipworm is from 

4 to 5 cm. long, % of its 
length consisting of a 
whip -like tail. It in- 
habits the cecum, but 
also the colon and rare- 
ly the small intestine. 
This is, perhaps, the 
most common of intes- 
tinal parasites since 
about 10.3% of adults 
in this country are in- 

FiG. 9-2. — Caudal bursa of Uncinaria duodenalis. (Schematic.) fected, but 45% in 

some parts of Germany and even 100% in Southern Italy. The eggs 
(Fig. 73) are very characteristic, from 50 to 54^ long and 23/x wide, 
with an unsegmented yolk and a very thick shell, into each pole of which 
is inserted a plug. We have found eggs which were certainly those of this 
worm which had no plugs. These eggs present an interesting variety of 
colors, some being light lemon-yellow, some deep yellow and some a dark 
brown! While harmless as a rule, this worm may cause enteritis and 
even a very severe, even fatal, anemia. Becker 23 classifies the symp- 

23 Deutsch. med. Wochenschr., June 26, 1902. 




THE INTESTINAL CONTENTS AND THE FECES 413 

toms of this infection as follows : gastro-intestinal symptoms among which 
are diarrhea due to ulcers or catarrh, blood in the stools and pain even sim- 
ulating appendicitis; nervous, some symptoms simulating meningitis 

(beriberi even has been blamed ... , — .. ,_ 

to this worm) and anemia, oc - J? £g ,: y^~ 

with all its symptoms '■ -. '" 

Strongyloides Stercora- t&.yQ 
lis; Anguillula Stercoralis 
et Intestinalis ; Leptodera . ' 
Stercoralis et Intestinal — /.'., v 
Rhabditis Stercoralis — . '* • v 

Rhabdomena Strongyloides, - : '- , 

etc — The rhabditiform larvae •- 

of strongyloides stercoralis ,■ - '■/• ' 
found in the stools measure . 
from 0.3 to 0.6 mm. long and I _ •" ;i'- ii-.jil.-gj'. ; ->^ * 

from 1 6 to 2?U wide Since ^ IG " 93 ' — Embryo of Uncinaria (americana?) found 

"^ ' in the stool. ^< 400. 

these embryos are very active 

in their motion the best way to find them is to make a depression in the fecal 
mass, fill it with water, then place the stool in a thermostat and the next day 
examine a drop of this water for these eel-like active worms. The eggs 
are very rarely present in the stools since practically all hatch in the bowel 
where all stages of the embryos may be found. If found, they could 
hardly be distinguished from those of Uncinaria duodenalis although 
they are perhaps a little larger (measuring from 65 to 70^ long and from 
34 to 39^ wide) and would contain a further developed embryo. 

The adult female resembles a filaria. It measures from 2.1 to 2.2 mm 
long and 32 to 39^ wide. The body increases slightly and gradually 
in size from the head to the posterior quarter, and then terminates 
rather suddenly in a short tail. The male is about % smaller. The 
worms are abundant in the duodenum and scanty in the jejunum. The 
adults are very rarely found in the stools. This infection was present in 
about 0.6% of a series of patients examined in Washington. Houghton, 
writing from Wuhu, China, states that from 0.9 to 1% of all patients ex- 
amined there harbored this parasite, but he emphasized the local distribu- 
tion of the parasite by saying that this worm has not yet been reported 
north pf the Yangtse River, nor in Manchuria and Korea. 

Trematodes. — Infections in the Far East by Schistosoma Japonicum 
have recently been carefully studied by Katsurada in Japan, by Catto in 
Singapore, by Woolley in the Philippine Islands and by Houghton 24 in 
China (in the provinces of Hunan, Honan, Hupeh, Kiangsi and Anhui). 

Houghton found that 8% of all male patients admitted during 1 year 

24 Trans, of the Society of Trop. Med. and Hygiene, June, 1910, vol. iii, No. 7, 
P- 342. 



414 CLINICAL DIAGNOSIS 

to the Wuhu General Hospital, Anhui, harbored this worm. Almost all 
of these were farmers and boatmen from the southern half of the province 
of Anhui and within a radius of ioo miles of Wuhu. The area of distribu- 
tion of this worm is sharply denned since Houghton writes (personal com- 
munication) that in the province adjoining Anhui on the North the worm 
has not yet been found. The distribution of the parasite in China follows 
in general the flat, low-lying lands of the central valley of the Yangtse and 
the valleys of tributary waters. In Anhui at least its presence is limited 
to the rice-growing divisions of the country. No cases from the hill or 
mountain districts have appeared. Peake 25 reports that it is common 
among raftsmen in Hunan. In some localities it is claimed that i in every 
3 or 4 or more of the farmers and boatmen shows the physical signs of this 
infection. 

Well-marked cases of this infection have enlarged liver and spleen, 
cachexia, eosinophilia, ascites, greatly exaggerated knee-jerks and bloody 
stools. The percentage of eosinophile cells in the blood varies from io to 
51% (average of 25%) of a total leucocyte count of from 2000 to 8500 
per c.mm. The anemia is seldom marked (Hb averages 80%). Less 
well-marked cases may have the enlarged spleen and the eosinophilia or the 
eosinophilia alone. Very few have ova in the stools as the only symptom. 

The ova may be found in the blood, but are more easily, demonstrated 
in the stools although it is not always easy to find them there, since their 
envelopes are sticky and so gather debris in the stool and leucocytes in 
the blood. They closely resemble the ova of Ascaris lumbricoides (they 
measure from 60 to 90// in length and 30 to 50^ in width), for which, under 
the low power, they may easily be mistaken, although they are more 
refractile, with envelopes not as deeply bile-stained and bosses not as 
prominent. These ova have a yellowish-brown color, are oval and have 
neither operculum nor spine. In the fresh stool the embryo, shaped like 
a melon seed, is quiescent in the egg but a little later the cilia are seen to 
be in motion. The free-swimming miradidium is found only after the 
stool has stood for about 10 hours. It can be kept alive in water for at 
least 5 days. 

The adult male measures about 10 mm. in length and 0.5 mm. in breadth. 

The slender, almost cylindrical female, is from 8 to 12 mm. long and 0.113 

mm. in diameter. The skin of this worm, unlike that of Schistosoma 

hematobium, is smooth. The adult worms are found in the smaller mesen- 

* 

teric blood-vessels (perhaps in the arteries especially) . The ova are found 
in necrotic areas in the mucosa and submucosa of both the small and the 
large bowel, but some are found in the subperitoneal tissue. 

Fasciolopsis buski (Distomum buski, D. crassum) the largest of the trematode 
parasites of man, measures from 34 to 70 mm. in length and from 5.5 to 15 mm. in width. 
Its eggs are from 120 to 130/1 long and from 77 to So/x wide. They have a thin shell, a 

25 China Medical Journal, 1908. 



THE INTESTINAL CONTENTS AND THE FECES 415 



very small operculum, and granular contents. Only a few cases, all of them intestinal 
infection, have been reported, and these were in the Far East. These patients suffered 
from a moderate diarrhea which continued for years, emaciation and anemia. The i 
parasite described under the name of Distomum rathouisi 
was, according to those in a position to judge, probably a 
specimen of Fasciolopsis buski. 26 

Distomum Lanceolatum. — The body of Distomum 
lanceolatum is pointed at both ends and measures from 8 to 
10 mm. long and from 1.5 to 2.5 mm. wide (see Fig. 94). The 
eggs, which are yellowish when fresh and dark brown later 
have thick shells and measure from 38 to 45/x in length and 
from 22 to 33/x in width. They contain an oval miracidium 
of which the anterior part alone is ciliated, and which hatches 
only in the intestine of some intermediary host, perhaps of a 
slug. This lancet fluke is a relatively rare parasite of the 
biliary ducts of the European and American domestic ani- 
mals. Thus far it has been found but seven times in man. 

Fasciola Hepatica. — The liver fluke is a widely- 
spread parasite inhabiting the bile-ducts of many herbivorous 
mammals. The adult measures from 20 to 30 mm. in length 
and from 8 to 13 mm. in breadth and has a definite head 
cone. The ova are yellowish brown in color, oval, measure 
from 130 to 145M long and from 70 to 90/x wide and have a 
cap-like lid. The elongated miracidium which is completely ciliated, escapes from 
the egg after this has been in the water for a few weeks and swims free until it 




a. c 

Fig. 94. — Dicrocelium lance- 
olatum, a, adult; b, egg with 
embryo; c, empty shell 
(From v. Jaksch.) 





Fig. 95. — A, ripe link of Tenia saginata. 



X 3. B, four unripe links. X 3. 



enters a water-snail in which host it passes through the stages of sporocyst, redia and 
cerecaria. The cerecaria become encysted on the grass of the meadows and is eaten by 
sheep, cattle, etc. Only 23 cases have been reported in man. 

Cestodes. — In a suspected case of tapeworm infection it is always 
important that segments be found before the treatment is undertaken. 

26 Jeffreys and Maxwell, Diseases of China, 1910. 



416 



CLINICAL DIAGNOSIS 





Pig. 96. — Eggs of Taenia sa- 
ginata. X 400. 

have been described. 



Mucus casts of the intestine, certain food constituents, etc., are often mis- 
taken for tapeworms. The treatment can be termed successful only if 
the head can be found (to find this, the stool is well mixed with water 
and allowed to settle for about 10 minutes, the heavy head will settle to 
the bottom, and the fluid is then decanted and this procedure repeated 
several times) or 3 months have passed with 
out the reappearance of segments. 

T^nia Solium (Figs. 97 and 98). — The infec- 
tion with Taenia solium is derived f rom Cysticercus 
cellulosae of pork. The adult worms in the intes- 
tine average about 3m. long, although much longer 
The head varies from 0.6 to 1 mm. in diameter 
and has four suckers from 0.4 to 0.5 mm. in diameter and a rostellum 
with a double crown of 22 to 32 
hooks from o . 1 1 t o o . 1 8 mm. long . y ■ n • ! ' ! 0T ^ ' ec 
The neck is about 3 cm. long, and is 
unsegmented. The ripe segments 
are from 9 to 10 mm. long by from 
4 to 5 mm. wide. The genital open- 
ings are marginal and alternate in 
a fairly regular manner. The shape 
of the uterus is one of the most 
characteristic points for identifi- 
cation. It consists of a large 
median stem with from 7 to 10 
coarse branches on each side, each 
of which branches dendritically. 
The eggs are round or oval, about 
3 5^ in diameter and the shell very 
thin and surrounded by a thick 
embryonic shell, radially marked 
and often yellow in color. This 
worm is very rare in America. 
The only specimen of Taenia sol- 
ium which we have seen (Figs. 97 
and 98) was discovered in Balti- 
more by Dr. Thos. Boggs. The 
most of those exhibited in mus- 
eums are wrongly labelled. 

T^nia Saginata. — The beef tapeworm, which infection is derived 
from the cysticercus of beef and, perhaps, of sheep, is quite common in 
this country. The adult worm varies from 4 to 8 m. or more in length. 
The head (Fig. 99) is from 1.5 to 2 mm. in diameter, is cuboid in shape and 
has 4 suckers, each 0.8 mm. in diameter and no hooks. The neck is long 



%H, 







Nat. size 



x4 




Fig. 97. — Tenia solium. Mature links from a case 

recently discovered in the Johns Hopkins Out-patient 

Department. (Kindness of Dr. T. R. 



THE INTESTINAL CONTENTS AND THE FECES 



417 




Fig. 98. — Taenia solium: Head (magnified), proglottis 
(actual size), and egg (magnified). (Zeiss's eye-piece 
IV, objective IV.) (From a preparation by Cori and 
v. Jaksch.) 



and delicate. The ripe segments (Fig. 95) are from. 16 to 20 mm. long and 
from 5 to 7 mm. wide. The over-ripe segments are longer and somewhat 
more slender. The genital openings are marginal and alternate irregularly. 
The uterus is characterized by the great number of its fine branches, from 
20 to 35 on each side of the median stem each of which branches dichoto- 
mously. The eggs (Fig. 96) are 
spherical or oval, from 30 to 40/x 
long and from 20 to 30^ wide and 
have a thin shell surrounded by 
an embryonic shell which is thick 
and radially striated. 

Hymenolepsis Nana— Taenia Na- 
na (Fig. 100). — This dwarf tapeworm is 
not at all uncommon in man, as Stiles 27 
has shown, who found it in 16 of 3500 
persons examined. 

The worm (Fig. 90) is from 10 to 
15 mm. long and from 0.5 to 0.7 mm. 
broad. Its spherical head, which is 
from 0.25 to 0.3 mm. in diameter, has 
4 suckers and a row of 24 to 30 very small and characteristically shaped hooks (14 to 
18//) long. The segments, about 150 in number are short (0.014 to 0.030 mm.) and 
relatively broad (0.4 to 0.9 mm.) 

The egg is characteristic in appearance. It is spherical or oval in shape from 30 
to 37 by 48/z in its 2 diameters and has 2 distinct thick membranes, the inner of which 
has at each pole a more or less conspicuous process with filamentous appendages. 

The parasite inhabits the 
ileum where a few or many 
thousands may be present. It is 
probably the same as the very 
common form in rats. 

DlPYLIDIUM CANINUM TE- 
NIA Cucumerina. — This tape- 
worm is from 15 to 35 cm. long 
and from 1.5 to 3 mm. broad. 
Its head is club-shaped with a 
rostellum and 3 or 4 rings of 
hooklets. The eggs are circular, 

from 43 to 50/-1 in diameter and the shell thin. It occurs in dogs, cats, and rarely in man, 

and then especially in children. 

Bothriocephalus Latus. — This tapeworm (Fig. 10 1), the largest 
parasite of man, is exceedingly common in the maritime countries of Europe, 
in Ireland and in Japan. A rapidly increasing number of cases is being 
discovered in America. The cysticercus stage occurs in fish. It often 
reaches 8 m. in length and a few even 50 feet. The infections are often 
multiple. In Willson's case 28 (this is 1 of the best reports) 2 worms were 

27 New York Med. Jour., 1903, vol. lxxviii, p. 877. 

28 Am. Jour, of Med. Sci., 1902, vol. exxiv. 




Fig. 99- — Head of Taenia saginata. X 5. 



418 



CLINICAL DIAGNOSIS 



present whose total length aggregate 82 feet. One man may harbor from 
50 td 100 of these worms but in such cases the individual worms are much 
shorter, perhaps from 3 to 5 feet long. The head is 1 mm. broad, from 
2 to 3 mm. long, is flat, almond or spoon-shaped, with 2 deep grooves at 
its sides-which serve as suckers. It has very little neck. The ripe segments, 
which begin about 50 c.c. from the head, increase in size until they reach 
from 10 to 15 mm. broad and 3 to 4 mm. long. The genital opening is on 

the side, not the edge, and 
around it the uterus radiates 
in the form of a rosette. The 
distribution of these organs 
is more regular than that of 
the septa of the segments. 
Willson considers that the 
presence of these imperfect 
or abortive segments is very 
characteristic of this family 
of worms. The eggs are char- 
acteristic. Their diameters 
are 70 and 45/*, their shell 
thin, the contents granular, 
giving them a mulberry-like 
appearance. The shell has a 
lid which may be open or 
closed. In very young eggs 
the lid is not evident. It 
may not be in older ones but 
can be rendered visible by 
pressure on the glass. The 
eggs are important in diag- 
nosis since the segments rare- 
ly appear in the stools, 
although when they do it is in 
great numbers. This tapeworm is important since some, a small percentage 
of its hosts, develop an anemia which hematologically cannot be distin- 
guished from primary pernicious anemia, and these recover rapidly after 
the worm has been expelled. Dehio thinks the worm, to produce this 
effect, must either die or at least be diseased and so furnishes a toxin 
which affects the bone marrow. 

The eggs of Schistosoma Hematobium (see page 418) also are met 
with in the stools of infected cases (Figs. 102 and 103). 

Plant Parasites. — Yeasts are often present in normal stools. Moulds 
are rare. Blastomycetes are found in the stools of patients with systemic 
infection with this parasite. 29 Oidium albicans has very rarely been found 
29 Fontaine, Hasse and Mitchell, Arch. Int. Med., Aug., 1909. 




Fig. 100. — Hymenolepis nana. Adult (left), head (right), 
egg (above); a, hooklet. (From Braun.) 



THE INTESTINAL CONTENTS AND THE FECES 



419 





in the stools of children. Sarcincz are often found in cases with dilated 
stomach and when present in large numbers may cause a diarrhea by the 
products of their fermentation. 

Bacteria. — The bodies of bacteria form a considerable part of the mass 
of the stools (see page 387) but the vast majority of these organisms aie 
dead. Almost any organism may gain entrance 
accidentally to the intestine, but there is a flora 
of bacteria so constant in the bowel that its 
presence may be considered normal. Among 
these are: Bacillus coli communis (see page 
286), Bacillus lactis aerogenes (see page 288), 
and for the suckling, Bacillus bifidus. 

Bacillus Bifidus 30 (Teissier, Paris Thesis, 
1900). — This organism (see Fig. 104) would 
appear to be a normal inhabitant of the intes- 
tines' of the suckling and to disappear soon Fig. 101.— Bothnocephaius latus. 
after the child is weaned. When, however, it 

persists, its presence would seem to be associated with symptoms of 
intestinal intoxication. It is an organism from 2 to 4^ long, and often 
in pairs. Its most characteristic shape is that of the letter Y. Involution 
forms are common. Its great interest is that this is 1 of the few intestinal 
organisms which are not decolorized by Gram's method. It is a strict 

anerobe. Among other important 
organisms are : Bacillus alkaligenes 
(see page 288) and the proteus 
group (seepage 288). Among the 
important pathogenic organisms 
which sometimes, even frequently, 
are found in the stools are: Ba- 
cillus pyocyaneous (see page 289); 
Bacillus aerogenes capsulatus (see 
page 289); Bacillus tetani (see 
page 290), the Staphylococcus 
group (see page 19) and the Strep- 
tococcus group (see page 19). 
A great many thermophilic and 
acidophilic organisms also are 
present which will not grow on ordinary media or at ordinary temperatures. 
The thermophilic organisms grow only at temperatures above 40 C. and 
some of them best at 6o° C. They, therefore, cannot multiply in the intes- 
tines and those found in the stools must have been swallowed with the 
food. The present opinion is that the lower bowel at least is not a favor- 
able habitat for organisms and that, as a result, the majority of those 
present in the stools are dead. 
30 Teissier, Paris Thesis, 1900. 



//J 

// / 


! 

1/ 








/. O C < \ V .:" Z> . 



Fig. 102. — Egg of Schistosoma haematobium found 
in stool. Embryo dead. X 400. 



420 



CLINICAL DIAGNOSIS 





Pig. 103.— Egg of Schistosoma 

haematobium found in stool. 

Embryo alive. X 400. 



Tubercle Bacilli. — In the search for Bacillus tuberculosis it is useless 
to try to digest a solid stool. Mucous masses, if present, should be selected, 
especially those which are blood-stained or purulent, and these are treated 
as is sputum. Indeed the diagnosis of pulmonary 
tuberculosis has frequently been made in this way, 
especially in children; but this is a rather remote 
possibility in the case of a careful adult. In intesti- 
nal tuberculosis these organisms are often present in 
abundance and yet in many such cases none at all are 
\ found, which has led to the supposition that they 
1 \ have been destroyed. 

Page's method of searching for Bacillus tubercu- 
^ J losis in the solid stool is to suspend a piece half the 
size of a pea in 1.5 c.c. of distilled water, add 54C.C. 
of a mixture of equal parts of absolute alcohol and 
ether and centrifugalize this for 10 minutes; a smear 
made of the sediment is fixed to the slide with a 
drop of egg albumin and stained as usual. 

To isolate tubercle bacilli from the feces 31 the 
specimen is diluted with about 3 volumes of water, 
well mixed and then filtered through several thicknesses of gauze to 
remove solid food particles. The filtrate is saturated with NaCl and left 
undisturbed for 30 minutes. The floating film (which contains all the 
bacteria) is then collected with a deflagration spoon in a wide-mouthed 
bottle and an equal volume of N NaOH ad- 
ded. This is shaken well and left for digestion 
in the incubator at 37 C. for 3 hours during 
which time it is shaken every half hour. It is 
then neutralized to sterile litmus paper, with 
N HC1, centrifugalized and the sediment inocu- 
lated into several test tubes of Petroff's 
medium. 

The growth appears in from 2 to 3 weeks, 
that is, much more slowly than in sputum 
cultures, probably since these bacilli have be- 
come weakened during their passage through 
the bowel. Many probably die which may 
explain the negative results in clearly tubercu- 
lous cases. 

Stools in Disease. — In typhoid fever the 
stool described as characteristic resembles " pea soup " in appearance, 
is copious, watery, of foul odor, alkaline in reaction and contains many 
triple phosphate crystals. Nevertheless in clinics which limit these 
31 Petroff, Jour. Exp. Med., 1915, xxli, 41. 




Fig. 104. — Bacillus bifidus. (Pho- 
tomicrograph by Dr. Thomas M. 
Wright.) 



THE INTESTINAL CONTENTS AND THE FECES 421 

patients to a careful diet such stools are rare and diarrhea is less com- 
mon than is constipation. The stool is frequently blood-tinged, this 
tinging sometimes warning us of a coming hemorrhage. Pus (microsco- 
pic) is rare except in severe cases with extensive ulceration. 

Bacillus Typhosus. — Many methods have been proposed for growing 
Bacillus typhosus from the stools. 32 

Drigalski and Conradi's medium 33 is the best. 

Three pounds of minced beef are mixed with 2 liters of water and allowed to stand 
over night. The beef is then pressed, the beef juice is boiled for 1 hour, filtered and to 
this filtrate are added 20 gms. of peptone (Witte), 20 gms. of nutrose and 10 gms. of 
sodium chloride. It is then boiled again for 1 hour and filtered. To this filtrate are 
added 60 gms. of the best agar. It is then boiled for 3 hours (1 of which is in the auto- 
clave). The fluid is made faintly alkaline to litmus, filtered and then boiled for % hour. 
While hot the litmus solution is added. The litmus solution is made by boiling 260 c.c. 
of litmus solution for 10 minutes, adding 30 gms. of the purest lactose and boiling again 
for 15 minutes. This litmus-lactose solution is now added while boiling to the hot agar 
solution described above. The mixture is then well shaken and then made very faintly 
alkaline. 

One then adds 4 c.c. of a hot sterile 10 % solution of soda and 20 c.c. of a freshly 
prepared 0.1 % solution of Krystallviolett B., Hochst, in warm sterile distilled water. 
The medium is poured at once into large Petri dishes or kept in 200 c.c. flasks. 

The resulting medium is a beef juice-nutrose-agar-lactose-litmus solu- 
tion which contains 0.01 p. M. per mille, i.e., per 1000 Krystallviolett. It 
hardens to a very firm, mass, firm enough to prevent much diffusion of any 
acid formed. It will not become dry. The lactose is split by Bacillus 
coli, not by Bacillus typhosus. The colonies of colon bacilli will therefore 
turn the medium red and the typhoid blue (since this organism splips off 
basic bodies from the proteids). Krystallviolett will inhibit the growth of 
many other organisms, especially acid-producing cocci. 

These authors recommend that several series of plates be inoculated in 
order to get the largest possible number of isolated colonies on a plate. 
If the stool is fluid 1 series of plates is inoculated with the undiluted stool 
and another with the stool diluted with 10 to 20 volumes of sterile normal 
salt solution. If the stool is solid it should be rubbed up to a homogeneous 
mass with sterile salt solution and various dilutions of this are used. 

The stool is rubbed onto the surface of the medium . After inoculation 
the plates are left open for at least half an* hour to allow the surface to dry, 
otherwise the colonies will coalesce. (The Krystallviolett will kill any air 
contaminations). When dry the plates are put into a thermostat at 37 C. 
and are examined in from 14 to 24 hours. The colon colonies will vary 
from 2 to 6 or more millimeters in diameter, are opaque, and will present 
a great variety of shades of red in color. The paratyphoid colonies are 
sometimes red and sometimes blue (see page 287). 

32 For a critical review of this subject, see Pratt, Boston, M. & S. Jour., 1907, vol. 516. 
33 Zeitschr. f. Hyg. u. inf. Krank, 1902, vol. xxxix, p. 283. 



422 CLINICAL DIAGNOSIS 

The typhoid colonies have a diameter of from i to 3 mm., are blue or 
violet in color, glassy, not doubly refractile and are seldom opaque. 

The colonies of the Bacillus subtilis group also are blue, but are so much 
larger than the typhoid that error is seldom possible. In some fetid stools 
the blue colonies of Proteus, Bacillus fluorescens and Bacillus fecalis alkal- 
igenes may deceive. They are rare and can be distinguished by the agglu- 
tination test. 

Peabody and Pratt 34 have shown the value of Malachite-green bouillon 
as an enriching medium. (The beef bouillon they used contained 1 : 1000 
malachite green and had an acidity of 0.5% to phenolphthalein. The 
amount of dye and the acidity must be determined for each preparation 
of the malachite green used.) This will completely inhibit the growth of 
Bacillus coli, but while Bacillus typhosus often will grow luxuriantly in it, 
ye,t the dye does exercise some restraint over this organism also. Tubes 
containing from 10 to 15 c.c. of this medium are inoculated with 1 drop of 
the fluid stool or suspension of the stool and are left in the thermostat for 
from 10 to 24 hours. Then 1 drop of the culture is rubbed over the surface 
of a Drigalski-Conradi plate. 

Drigalski and Conradi were able to grow typhoid bacilli from the stools 
of every case of typhoid fever they examined. Pratt and Peabody 35 were 
able to find it in but 2 1% of their cases. These bacilli were most numerous 
in those stools which contained blood. Many believe that while it is pos- 
sible that all patients with typhoid fever may have a few living typhoid 
bacilli in their stools yet that the great majority are destroyed in the intes- 
tine and that only a few are alive when the stool reaches the rectum. 

In severe Asiatic cholera the rice-water stools are quite character- 
istic. They are copious and consist chiefly of water secreted by the intes- 
tinal wall in which float small gray flecks which are masses of epithelial 
cells, cholera spirillar and fat droplets. They have no fecal odor, are alkaline, 
almost acholic, sometimes are blood-stained and contain little albumin 
and much salt. 

Spirillum Cholera Asiatics. — The spirillum of Asiatic cholera, 
or the " comma bacillus," is a small curved " comma- shaped " bacillus, 
about 2/jl long and 0.5/* thick. It is very actively motile. It has a single, 
long, delicate flagellum at one end, it does not produce spores, it is readily 
stained in all bacterial stains and it decolorizes by Gram's method. Involu- 
tion forms are common. This organism is very aerobic, a very rapid grower 
at room temperature on all ordinary media and also in some which are so 
poor in nutriment that other organisms cannot multiply at all in them. It 
will not grow on potato at room temperature, but will if in a thermostat. 
Its growth on gelatin, which it liquefies, is fairly characteristic in that the 
colonies soon appear as minute white points which resemble fragments of 

34 Bogton MH. and Surg. Jour., Feb. 13, 1908. 
36 Jour. Am. Med. Assn., 1907, xlix, 846. 



THE INTESTINAL CONTENTS AND THE FECES 423 

broken ground glass with granular irregular margins. Later, liquefaction 
begins and the colony sinks in the little cup of liquid cloudy gelatin which 
forms a halo around it. This organism produces much indol. It is very 
sensitive to acids. 

The non-pathogenic spirilla are very common. More than 60 species 
with similar morphology, but different cultural characteristics, have been 
found in various drinking waters (e.g., Spirillum Schuylkiliensis) , while 
there is 1 in the mouth which will not grow at all in ordinary media. 

Many pathogenic forms also have been described: Metchnikoff's spiril- 
lum, found in an epidemic in chickens, is pathogenic for birds while the 
true cholera spirillum is not, and is a more rapid grower than is the latter; 
Massea's spirillum is very pathogenic to pigeons and has 4 or 5 flagella; 
Finkler and Prior's form, from a case of cholera nostras, will grow as a 
dirty brown scum on potato at room temperature; while Deneke's form, 
from old cheese, will not grow on potato. 

But their morphology and cultural characteristics are not sufficient 
for the recognition of these organisms. The following specific biological 
test also must be used. A guinea-pig is first immunized to 1 species of 
spirillum and a small dose of the organism to be examined is then injected 
into its peritoneal cavity. If the organism injected is the 1 to which this 
animal has been immunized those introduced will be rapidly destroyed. 

The diagnosis of Asiatic cholera often may (in some epidemics from 25 
to 50% of the cases) be made directly from the stools, a smear of which 
will show vast numbers of these comma-bacilli. Usually, however, it is 
necessary to inoculate gelatin and agar plates with " rice particles " from 
the stool. The typical colonies will be recognized in 24 hours if grown on 
gelatin at 22 C. When but few organisms are present the enriching method 
of Schottelius is of value. That is, one incubates a large amount of bouillon 
with a little of the stool and inculcates this for a few hours, in which time 
these spirillae will form a surface scum from which cultures may be made. 
This method' is of value in the study of drinking water. To 90 cc. of the 
suspected water are added 10 cc. of a sterile solution of 10% peptone and 
5% of sodium chloride. This is incubated in a thermostat and the scum 
later examined. 

In DYSENTERY, RECTAL DIARRHEA, and CANCER OP THE RECTUM the 

movements are frequent and scanty. They soon lose their fecal character 
and become mucoid, mucopurulent, hemorrhagic or sero-hemorrhagic. 
According to the amount of blood present these cases have been classified 
as " white " and " red " diarrhea. Among the masses of bloody mucus 
may sometimes be found fragments of necrotic mucous membrane or of 
cancer. In amebic dysentery, during the acute exacerbations, a diarrhea of 
from 3 to 6 movements a day is the rule. The stools are thin and watery, 
offensive in odor, and contain bloody mucus in which the amebas may be 
found. These periods of diarrhea are separated by even years during 



424 CLINICAL DIAGNOSIS 

which time the movements are normal or constipated; and yet even in 
these stools the ameba may be found. It is for this reason that a routine 
examination of the stools for ameba should be made even in cases without 
dysentery or symptoms referable to the liver. (This was well illustrated 
by a case of constipation with irregular fever and without symptoms of 
hepatic or intestinal trouble. The autopsy revealed a large amebic abscess 
of the liver. 36 ) 

The Group of Dysentery Bacilli. — In morphology and in some 
cultural characteristics " Shiga's bacillus " resembles Bacillus typhosus. 
It is a short organism with rounded ends and is inclined to involution 
forms. All now agree that it is non-motile. No spores are formed. This 
organism stains readily in the commonly used aniline dyes, shows a tend- 
ency to polar staining and is decolorized by Gram's method. 

Since Shiga described his organism 12 other organisms of dysentery 
have been described, all belonging to 1 group, all with similar morphology 
and similar staining characteristics, all non-motile, all unable to liquefy 
gelatin, to form acid from lactose and to form gas from any carbohydrate. 
They differ in their agglutination reactions to immune sera and in their 
ability to ferment carbohydrates. Flexner recognizes 3 types : 1 . The 
" Shiga " type, which ferments glucose only. 2. The " Flexner-Harris " 
type, which ferments glucose, mannite and dextrin, but not lactose. 
This is the type which prevails in the United States. 3. Bacillus " Y ,; 
(His and Russell), which ferments only glucose and mannite. 

These organisms cause the so-called " bacillary " or " infectious " 
dysenteries, diseases which may occur sporadically or in epidemics, e.g. the 
severe epidemics of tropical dysentery. This disease begins as an acute 
gastro-enteritis with a diarrhea which increases in severity until the stools 
are very frequent and scanty, lose their fecal character and consist chiefly 
of mucus and blood rich in these organisms. 

In the recognition of the dysentery bacilli the agglutination tests are 
of greatest value. The blood serum of a patient infected with an organism 
belonging to the Flexner-Harris type will agglutinate a pure culture of this 
organism in dilutions of 1 : 1000-1500. In cases of infection with one of 
the Shiga type the agglutination is less complete. 

Pancreatic Disease. — The pancreas may be diseased in part or as a 
whole. Chronic glycosuria is ascribed to a lesion of the islands of Langer- 
hans. In these cases there is often no evidence of disease of the rest of this 
organ. In cancer of the pancreas or total obstruction of the duct from cal- 
culus leading to atrophy of this organ, the pancreas as a whole is destroyed, 
but first the tissue which produces its external secretion. There is no 
quantitative relation evident between the lesion and the impairment of 
pancreatic function, for the first evidence of pancreatic disturbance may 
be late, the glandular insufficiency developing all at once as it were. There 

36 See also Councilman and Lafleur, Johns Hopkins Hosp. Reports, vol. 11, p. 395. 



THE INTESTINAL CONTENTS AND THE FECES 425 

are no tests for partial pancreatic insufficiency, which have any proven 
practical, or even theoretical, value. 

The failure of internal secretion has been discussed on page 198. The 
3 signs of entire lack of external secretion are steatorrhea, azotorrhea and 
the lack of the products of putrefaction in a case without diarrhea and 
without jaundice. 

By azotorrhea is meant the presence in the stool of a person on mixed 
diet who has no diarrhea of an unusual number of muscle fibers, some still 
in bundles, with their striation well preserved. 

Steatorrhea. — In some cases of diabetes the stools contain from a few 
drachms to a cupful of pure yellowish-brown fat. Others consist of about 
30% fat. The suspicion of pancreatic disease is certainly justified if a large 
amount of fluid fat separates itself from the rest of the stool. The simul- 
taneous presence of glycosuria in such cases is rare, the absence of decom- 
position is not usual nor should it be expected since so much albumin is 
present, but a simultaneous steatorrhea and azotorrhea are important 
and with diabetes are conclusive. 

And yet azotorrhea may wholly fail in pancreatic disease. Again, stea- 
torrhea alone is not conclusive for with complete atrophy oi the pancreas 
steatorrhea may fail, while it may be present in many diseases other than 
of the pancreas which affect fat absorption as well as fat splitting. Again, 
in some cases with an increased amount of fat in the stool the per cent, 
split is normal. 

Atkinson and Hirsh 37 reported a typical case of severe chronic inter- 
stitial pancreatitis due to pancreatic lithiasis. The patient, one of diabetes 
mellitus, evacuated 4 liters of feces daily. The stools were soft or semi- 
solid, leathery-brown in color and had the odor of rancid butter, they 
contained 54.6% of fat (22.5% neutral fat and 32.1% fatty acid), which 
was present in lumps varying in size from that of a split pea to that of 
a walnut. 

The assimilation limit for fat for a normal person is about 350 gms. of 
butter. After meals not exceeding this amount the loss in the stools is not 
over 7 to 10%. This test should not be made if the person is jaundiced 
or has acholia due to any other disease; the fat should not be given in an 
emulsified condition and any diarrhea should be checked with opium prior 
to making this test. 

The attempt has frequently been made to find a preparation of pancreas 
which will not be affected by the gastric juice. If the administration of 
such a preparation to a case without diarrhea and with considerable muscle 
tissue in the stool were followed by a diminution of the muscle fibers, the 
evidence would be in favor of pancreatic disease. These attempts are 
partially successful if the gastric secretion be kept alkaline after the inges- 
tion of the pancreon, pancreatin, trypsin, etc. 

37 Am. Jour. Med. Sci., Oct., 1907. 



426 CLINICAL DIAGNOSIS 

Permanent Mounts of Small Worms — For the following methods I 
am indebted to Dr. Thomas R. Boggs. 

Boggs' Method. — The worm is allowed to die in water (that it may be 
fixed while in a relaxed condition). It is then spread out on a piece of 
filter paper and immersed in an alcohol-glycerin cleaning fluid (alcohol 
80%, 16 parts, and glycerin 4 parts). The specimen is allowed to remain 
in this solution in an open dish loosely covered with cloth or paper, until 
the alcohol has entirely evaporated off. This may take from 2 to 6 weeks. 
Since the worm is spread out on the filter paper it will contract but little. 
Should it do so it may be slightly pressed between slides held together by 
rubber bands. When the specimen is sufficiently clear it is gently blotted 
on the slide and covered with melted glycerin jelly. The cover slip is then 
dropped on and if necessary pressed down until the jelly has hardened. 
After the jelly is hard the excess is removed from the borders of the cover 
and the edges sealed with microscope cement or asphalt paint. 

The glycerin jelly is made by melting 14 gms. of the best gold mark 
gelatin in 120 c.c. of hot water and adding 120 c.c. of glycerin. This is then 
cooled to 50 C, the carefully separated whites of 2 eggs added, the fluid 
heatedgently without stirring and then filtered. The volume is now madeupto 
240 c.c. by adding water and 1 c.c. of pure carbolic acid added. This jelly is 
solid at ordinary temperature, but is easily melted under the hot water tap. 

To Preserve Stools Containing Parasite Eggs. — The stools 
are diluted to a soup-like consistency and then #o its volume of formalin 
added. The specimens are then kept in a tightly corked bottle. Parasite 
eggs and larvas will be well preserved. 

Flat Worms, Preservation of the Gross Specimen. 38 — To clean 
the worms the fecal matter is mixed with warm (37 to 40 ) normal salt 
solution sufficient to make a thin broth. If the specimens are obtained at 
autopsy, the intestinal contents may be washed or scraped off into the salt 
solution. The worms will move about freely and are easily seen and iso- 
lated, especially small worms, such as Hymenolepis nana. With a pair of 
finely-pointed forceps the parasites are picked up and transferred to a 
second dish of warm solution. Specimens to be cut in sections may be 
taken from this solution clean and treated with the proper fixatives (see 
below). The rest of the material may be placed in 50 to 70% alcohol 
with or without glycerin, or in Zenker's solution, or in a 2% solution of 
formalin. Zenker's solution causes considerable shrinking and a rather 
marked yellowish discoloration. These authors consider the formalin 
mixture much better as it preserves the natural whiteness of the worms 
and causes little or no shrinkage. 

Preparation of segments for mounting. — The specimens are washed in 
normal salt solution (0.85%) and fixed by keeping them from 14 to 16 hours 

38 Mink and Ebeling, U. S. Naval Med. Bull., No. 3, vol. 111. 



THE INTESTINAL CONTENTS AND THE FECES 427 

in a 2% formalin. They are then transferred to the following glucose 
medium, which is a slight modification of the Fabre-Domerque medium : 

Sirup (glucose 48 parts; water 52 parts) 1000 c.c. 

Methyl alcohol 200 c.c. 

Glycerin 100 c.c. 

Camphor (q. s. to keep) 100 c.c. 

The specimens will clear sufficiently in this medium in 4 or 5 hours. 
They may, however, be left in it indefinitely. To mount them, a sufficient 
quantity of glycerin jelly is dropped on a slide and the specimen is trans- 
ferred to this, care being taken not to admit air-bubbles. A cover-glass 
which has been passed through the flame finishes the mount. After the 
glycerin jelly has hardened a few coats of gold-size applied around the 
cover-glass will furnish rigidity and improve the general appearance of the 
preparation. Concave slides are desirable when the specimen is of uneven 
thickness or rather thick throughout. 

Preparation and sectioning of material. — To prevent any curling or 
distortion of the worm or segment a fixative should be used which will 
kill quickly. For this Zenker's fluid is the best. About 3 or 4 inches of 
the live tape-worm are taken from the salt solution and stretched out on 
an ordinary glass slide. By means of a pipet the slide is rapidly covered 
with Zenker's fluid. The worm will straighten out, harden, and float on 
the solution. It may then be transferred to a flat dish filled with Zenker's 
fluid where it should remain for from 2 to 24 hours. By cutting slightly 
beyond the part needed for work one leaves small end pieces which may be 
grasped with the forceps in the subsequent manipulations and so the 
segments chosen for study will not be touched by the forceps. The later 
processes includes treatment in an alcohol-iodine solution, in graded alco- 
hols for dehydrating, and other steps until the tissue is immersed in melted 
45 C. paraffin. In blocking the specimen it seems best to place the longest 
and broadest surface downward and later trim and mount it as desired. 
Specimens are best cut either in planes parallel with the long, broad surface, 
or perpendicular to the long axis. They should be from 25 to 30/* thick. 
The most convenient stain is a rapidly acting, purely nuclear hematoxylin. 

vStained specimens of worms. — The worm, as fresh as possible, is fixed 
in a boiling, saturated alcoholic solution of mercuric chloride for from 10 
to 30 minutes (depending on the thickness of the specimen). It is then 
washed over night in running water and then in water containing a trace 
of iodine until it is free from mercury. The specimen is then heavily over- 
stained (for from 12 to 24 hours) with hematoxylin or carmine and decolor- 
ized under the lower power of the microscope with acid alcohol until the 
desired color is obtained. The specimen is then washed, dehydrated in 
alcohol, cleared in oil of cloves or creosote and mounted in Canada balsam. 
This method is best for the study of the minute anatomy of tapeworms 
and flukes. It is not successful with round worms. 



CHAPTER V 

THE BLOOD 

Instruments for Obtaining the Blood. — If a few drops of blood will 
suffice for examination one usually pricks the lobule of the ear or the tip of 
a finger, using a simple sharp-pointed lancet, a needle with a cutting edge 
or an ordinary Hagedorn needle. Special forms of lancet have been in- 
vented, some, as the Daland needle, with a guard which prevents the needle 
from penetrating beyond a certain depth, others with a spring which when 
released forces the needle to a certain depth (e.g. Francke's needle). 

If considerable blood is desired a hypodermic syringe should be used to 

penetrate into the lumen of a vein, 
preferably the median basilic vein at 
the elbow. The skin of this region is 
first made surgically clean, e.g., with 
tincture of iodine, then a tight band- 
age is tied around the upper arm and 
a towel wet with warm bichloride 
wrapped around the elbow- joint until 
the needle is inserted. If the blood 
is intended for quantitative work the 
bandage should be removed after the 
needle has been inserted and circu- 
lation allowed to return to normal be- 
fore any blood is withdrawn, since stasis will alter its concentration. 

Two forms of forceps are necessary in making blood smears (Fig. 105). 
One has crossed blades which will hold the entire length of one edge of a 
cover-glass firmly and the second, the ordinary straight pinch forceps, has 
flat smooth points and a weak spring. Cover-glasses should always be 
handled with forceps since the moisture from the fingers will affect the 
specimens. 

The best glass slides for blood work are of clear glass 1 inch wide and 
as flat as possible. Those which are slightly curved will rock on the micro- 
scope stage unless the specimen is on the convex side. 

The cover-glasses should be thin (No. 1, or preferably No. o) and not 
over % of an inch square. In general only new cover-glasses should be em- 
ployed, since it is almost impossible to remove from those once used the 
little microscopic masses of hemoglobin which latter may be mistaken for 
malarial pigment, etc. 

The glassware must be scrupulously clean. New slides and cover-glasses 
may be washed in soap and water, then in clean water and lastly in 95% 
alcohol. Some are so oily that they should be soaked for about 24 hours 
in concentrated hydrochloric acid, then washed in water, then in 95% 
alcohol and lastly in ether. The clean glass may be kept either in 95% 




Fig. 105.- 



•Method of making cover-glass 
preparations. 



428 



THE BLOOD 429 

alcohol or, carefully wiped (best with an old linen handkerchief) , in a dry 
glass dish. They should be handled only with forceps. 

The skin punctured for the blood should not be cyanosed or edematous. 
We have seen 2 leucocyte counts of blood taken at the same time from the 
2 ears of the same person vary by 100% and the same may be said of blood 
of the 2 hands. The blood is usually obtained from the ear since this is 
always within reach, the patient cannot watch the worker and the skin 
of the lobule is relatively painless. If the lobe of the ear is thick it is usually 
stretched over the index-finger by the thumb and middle finger and pricked 
on its fiat side, but if thin, and especially if several drops of blood are re- 
quired, it is well to pierce its edge. Many workers prick the palmar surface 
of the ball of a finger of the left hand where they easily obtain a good drop 
of blood. Our students studying their own blood search on the anterior 
surface of the forearm for the pain points and, avoiding these, prick over 
a small superficial vein. In this way a large drop of blood is easily and 
painlessly obtained. In the case of very small children the great toe or the 
heel is chosen. The needle or lancet, washed in soap and water, may be 
sterilized by dipping it into alcohol. The skin is washed off with alcohol 
and allowed to dry. The lancet may be driven through the skin by a short, 
quick, sharp blow; or if several drops are desired, by slow pressure. The 
patients much prefer 1 hard stab to several ineffectual ones. The skin once 
pierced should not be squeezed, nor rubbed, nor held in a position which 
will check its circulation since all these methods to increase the flow will 
change the concentration of the blood. The drops should well out. The 
first is wiped off and the second used. In case many drops are to be taken 
the incision may be wiped occasionally with an alcohol sponge and then 
with a dry sponge to keep the cut bleeding. Always in advance one should 
ask for a history of hemophilia and thus avoid hemorrhages difficult to 
check. In these cases the very slightest prick will furnish even too much 
blood, although even in these cases blood may be drawn from a vein pro- 
vided the vein is not wounded {i.e., the first prick successful). 

Specific Gravity of Blood — Gravimetric Method. — The gravimetric 
method of determining the specific gravity of the blood is certainly the 
most accurate, but requires at least 5 c.c. of blood for an accurate estima- 
tion, a good pycnometer and a very accurate chemical balance. 

Aremetrical Methods. — A popular method (Roy) since easy, is to 
introduce single drops of blood into a series of bottles filled with fluids 
previously prepared with which the blood will not mix and each of a differ- 
ent specific gravity and noting the one in which the drop of blood neither 
sinks nor rises. 

Hammerschlag changes a mixture of benzol and chloroform until it 
has just the right specific gravity. A glass cylinder, perfectly clean and 
dry (else the blood will cling to the side of the glass) , is filled with the mix- 
ture mentioned above, the specific gravity of which is about 1.058. A drop 



430 CLINICAL DIAGNOSIS 

of blood is then introduced, best through a capillary tube bent at the end 
at right angles so that the drop may be blown into the fluid without impart- 
ing to it any upward or downward motion. If the drop rises more benzol 
is added; if it sinks, more chloroform. After each addition the fluid must 
be well stirred. It is important to work very rapidly since the specific 
gravity of the mixture is constantly changing because of evaporation and 
since there is some exchange between the blood and the fluid which alters 
the drop of blood. The final result should be confirmed using a fresh drop 
of blood and working very rapidly. The drop of blood is removed before 
the specific gravity of the fluid is tested by filtering it through linen. Care 
must be taken that no bubbles of air are sticking to the drop. Slight differ- 
ences in the temperature of the mixture make differences in the specific 
gravity of the mixture which are so great that Langlois varies, not the 
proportions of benzol and chloroform in the mixture, but its temperature. 
When the drop no longer rises or sinks he reads the temperature of the 
mixture and from this reckons its specific gravity. 

The specific gravity of the blood serum (obtained by filling a tube with 
blood, sealing both its ends and allowing it to stand upright until the serum 
has separated well from the clot) and of the plasma (obtained by centri- 
f ugalizing the blood in a centrifuge tube which has been previously washed 
out with 3% oxalic acid) is estimated in a similar manner. 

While the Hammerschlag method looks easy and is simple the possi- 
bilities of error are so great that only one with considerable training can 
use it safely. 

The specific gravity of normal blood varies from 1.058 to 1.062 the 
average for men being 1.059 an d for women 1.056 (Ehrlich). The figures 
given by Piper are, for man 1.055, for woman 1.053 an d for children 1.05 1 : 
Landois, 1.045 "to 1.075, "the average 1.054: Lloyd Jones, 1.036 to 1.068: 
and Hammerschlag, from 1.056 to 1.063. Most agree that the specific 
gravity of the blood of a woman is slightly less than that of a man. That 
of the blood at birth is 1.066 (Lloyd Jones). It then drops, reaching a 
minimum of 1.048 to 1.050 in the second year and then rises to a maximum 
of almost 1.058 (even 1.066). After the menopause the average is 1.054 
Diet affects if but little. Menstruation, Schmalz says, is followed by a 
slight increase. Daily variations are noted by Schmalz, the maximum 
between 7 and 8 a.m. being 1.060—7, an d the minimum from 11 a.m. to 
8 p.m. 1.0588. 

The specific gravity for the serum and the plasma are about the same, 
from 1.029 to 1.032, average 1.030. The specific gravity of the plasma, 
while much more uniform than that of the total blood, nevertheless is 
diminished in dropsical conditions. 

Using the Hammerschlag method, 23 of our students, normal men between 20 and 
25 years of age, found their blood to vary from 1.051 to 1.065. In the case of 16 of the 
23 it was from 1.057 to 1.061 ; the mean of all was 1.058. 



THE BLOOD 



431 



In pathological conditions the specific gravity of the blood may vary 
from 1.025 to 1.068, running parallel in most cases to the hemoglobin. It 
is reduced in all anemias, especially in chlorosis. It is reduced in many 
cachexias, in which cases the change is in the plasma since the hemoglobin 
may remain practically normal. It is increased in fevers to from 1.057 to 
1.063, i n cyanosis and in obstructive jaundice. 

Until the introduction of the better forms of hemoglobinometer the 
hemoglobin was best calculated from the specific gravity. This was 
especially true of such anemias as chlorosis, in which cases variations in 
the specific gravity are almost entirely due to variations in the amount of 
hemoglobin. In cases with hydremia, however, this rule does not hold 
and yet even in these cases the specific gravity of the plasma is more con- 
stant than that of the total blood since the water would seem to be taken 
up in large part by the red blood-cells. This is true even in severe blood 
diseases, as, for example, in pernicious anemia. 

Dried Residue — Hygrometry. — To calculate the dried residue of the 
blood a weighing glass with ground-glass stopper is first carefully dried 
and weighed. A little blood is then introduced, the cover put in place 
and it is again weighed. The cover is now tilted and the blood dried to 
constant weight (about 24 hours) in a thermostat at a temperature of from 
65 to 70 C. It is then again weighed. The solids of the blood of a normal 
man average about 21.6% of the weight of the fresh blood; of a woman, 
19.8%. The figures of Askanazy are: for man, from 20.35 to 22.89%; 
average, 21.92%; for woman, from 19.58 to 21.46%; average, 20.53%. 

It was hoped that this study of dried residue would help in the study of 
the anemias, since it was found to vary somewhat independently of the 
specific gravity of the blood, of the red cell count and of the hemoglobin ; 
but its value has not yet been proven. 

Sedimentation of the Blood. — The attempt has been made to substitute 
the estimation of the volume of the red blood-corpuscles for cell counting, 
since after all it is not so much the number of the red blood-cells as the 
volume of hemoglobin-containing protoplasm which is important in internal 
respiration. While this substitution was not successful yet the sedimenta- 
tion test did win an independent value for itself (see page xxx). The 
volume of these cells may be determined by the hematocrit (see Fig. 122) 
using undiluted blood (see page 460), or by the centrifuge using blood 
diluted with an equal volume of 2.5% potassium bichromate or of Muller's 
fluid, or, most accurate of all, by the spontaneous sedimentation of the red 
blood-cells. The difficulty with the method is the varying compressibility 
of the red cells in different conditions. 

Normally the volume of these cells is 50% of the blood volume. 

Coagulation. — On few subjects in hematology has so much accurate, 
careful scientific work been done as on the coagulation of the blood and the 
results are of unquestionable clinical value. The subject is a difficult one. 



432 CLINICAL DIAGNOSIS 

There are at least 3 forms of coagulation to consider — that in an open 
wound, thrombus formation in a closed vessel and coagulation within our 
laboratory instruments. These processes are very different, and " we can- 
not bring the appearance of coagulation in the living vessel into direct 
parallelism with coagulation of blood as ordinarily understood " (Welch), 
nor can we reproduce the conditions under which either occurs. Throm- 
bosis is a very common complication in'typhoid fever, anemia and cachexia, 
yet the amount of fibrin demonstrable in the blood in these conditions is 
quite low; on the other hand the blood in pneumonia and in acute articular 
rheumatism is very rich in visible fibrin and yet thrombosis is of rare occur- 
rence. Again, coagulation in the wound is not a uniform process. More 
depends on the nature of the vessels cut and on the tissue through which 
the blood escapes. It is, for illustration, scarcely possible for a man to 
bleed to death following cross-section of a radial artery, large as that is, 
while the fatal intestinal hemorrhages in typhoid fever are from vessels 
so small that they cannot be found without a dissecting microscope. The 
character of the vessel's wall, the opportunity for it to contract, the charac- 
ter of the tissue of the Peyer's patch and possibly the intestinal contents — all 
may conspire to make the intestinal hemorrhage much more serious. Then 
too, the rapidity of coagulation of blood in a tube or glass chamber depends 
on many factors, few of which are understood. For instance, the longer 
the blood is in contact with the cut tissues the more rapidly will it clot; 
blood from a deep wound will clot even 3 minutes more slowly than will 
that from a superficial wound; each drop of blood flowing from a wound 
will clot more quickly than will the preceding drops so that if several are 
allowed to flow the difference in coagulation time between the first and 
last drop may be almost 10 minutes; the pressure made on the flesh near 
the cut to encourage the flow of blood, the amount of blood used, the 
material composing the receptacle in which it clots, this receptacle's clean- 
liness and temperature, the temperature of the air, and the. opportunity 
there is for evaporation — all these modify the rapidity of coagulation. The 
above considerations show the need there is of as uniform technic as possible. 

Even when the technic is as uniform as possible it is still true that blood 
removed at the same time from different parts of the body will clot with 
different rapidity; that the diet and especially the medicines are factors 
to be considered ; and that the coagulation time differs appreciably at differ- 
ent times of the day. The longest time noted is soon after breakfast, when 
the blood of normal men sometimes clots in from 12 to 17 minutes, a slow- 
ness which at any other time of day would be distinctly pathological. We 
wonder if those surgeons who prefer the forenoon hours for their operations 
remember this? The most rapid coagulation occurs about 4 o'clock in 
the afternoon. 

While we have not yet been able to gain much, if any, data concerning 
internal coagulation, as in thrombosis (save the importance of infection), 



THE BLOOD 433 

the clotting of effusions, etc., yet the observations made on cases with 
" the hemorrhagic diatheses," as the purpuras, hemophilias and the angio- 
neurotic edemas are of interest. Some surgeons appreciate the importance 
of the coagulation time in hemophilia and jaundice but many " still take 
a chance." 

A normal coagulation time has chiefly negative value; that is, if it is 
normal one may feel safe in operating, while if definitely slower than the 
limits of normal there may or there may not be danger. Hinman and 
Sladen l give the following illustration of this point : The coagulation time 
in a case of hemophilia was 16% minutes; that is, it was distinctly prolonged 
and yet the prick in the ear by which this drop was obtained closed at once 
without further bleeding. On the same day this same ear was pricked a 
second time. This drop clotted in 18)3 minutes but this prick bled for 
12 hours. 

Hemophilia is defined by Howell 2 as a condition limited to the male, 
the characteristic peculiarity of which is that the coagulation time of the 
blood is markedly prolonged in consequence of a deficiency in the amount 
of the contained prothrombin, with the additional characteristic that the 
defect is transmissable by heredity in accordance with the so-called law 
of Nasse. Since the prothrombin present in the plasma is furnished by the 
blood-platelets Howell considers it reasonable to assume that the defect 
in hemophilia is referable to some functional change in these elements. 
The antithrombin, which is diminished in cases of thrombosis, is relatively 
increased in hemophilia owing to the absolute decrease of prothrombin. 

In purpura hemorrhagica and other forms of so-called purpura no evi- 
dence was found of any variation from normal in either the antithrombin 
or the prothrombin. 3 

Estimation of Coagulation Time. — " Coagulation time " is not the 
time it takes blood to clot in a wound, for we have no means of measuring 
'that, but the time which elapses between the appearance in the wound of 
the drop of blood to be tested and the first evidence of fibrin formation 
in this blood in the laboratory instrument. One does not measure the time 
from the moment he gets it in the instrument. For comparable work if 
drops of blood are to be used all determinations should be made approxi- 
mately at the same time of day ; the blood should always be obtained from 
the same part of the body ; one cannot expect to get uniform flow of blood 
but he should at least get a free flow ; the prick should not be made in the 
seat of an active or a passive congestion (for tissue lymph and carbon 
dioxide both hasten coagulation) ; the second or third drops which well 
out should be used, not the first and not the later drops; the temperature 
of the room in which the observation is made should not be unusually 

1 Johns Hopkins Hosp. Bull, July 1907, xviii, p. 207. 

2 Arch, of Int. Med., 1914, xiii, p. 92. 

3 For the methods of determining the amounts of antithrombin and prothrombin 
see Dr. Howell's article. 

28 



434 CLINICAL DIAGNOSIS 

warm or cold, although one need not try to control the temper ature beyond 
preventing extremes; and anything which tends to increase the drying of 
the blood, as a draught of air, should be avoided. In the newer methods 
from 2 to 8 c.c. drawn from a superficial vein are used. 

The method used should allow the observer to prove objectively the 
presence of fibrin since the drop of blood may dry and appear clotted before 
any fibrin has formed. 

Vierordt's Method. — This method has simplicity to recommend it. A 
white horse-hair io cm. long is boiled in alcohol and ether. A capillary 
tube 5 cm. long and of i mm. bore is thoroughly washed and dried also in 
alcohol and ether. A drop of blood giving a column about 5 mm. long is 
received into the tube and the white horse-hair run through it. Each 
minute the hair is pulled slightly through the drop. The first appearance 
of coagulation is shown by a slight reddish stain on the hair, which after 
the blood is well coagulated will again appear clean. It is of greatest im- 
portance that that part of the horse-hair which is to come into contact 
with the blood should not ha\^e been touched with the fingers. The amount 
of blood used should be exactly the same each time, since the coagulation 
time depends directly upon the amount of blood. All results should be 
confirmed by a second determination. 

Millians method is a modification of Hayem's. This method which is 
considerably in vogue among the French is to place a drop of blood on a 
clean glass slide, cover it by a crystallizing dish to prevent very much 
evaporation and then at stated intervals to tilt the slide. From the change 
in shape of the drop of blood, that is, when it ceases to act as a fluid but 
has elasticity of form, can be determined the coagulation point. Most 
unreasonable results have been obtained by this method, the coagulation 
time extending even into hours. The method has been tested under Dr. 
Boggs' direction in the Johns Hopkins clinic by Hinman and Sladen 4 who 
found that very much depends on the size of the drop, which should be 
constant, and on the amount of evaporation. And yet if no better appara- 
tus is at hand this simple method has some value. 

McGowari 's Method. 5 — Capillary tubes of uniform caliber are filled with 
blood and small sections broken off at intervals of from 10 to 30 seconds 
until a fine filament of fibrin is observed between the carefully separated 
ends of the tube and fragment. 

The best method is that of Russell and Brodie. Their apparatus con- 
sists of a moist chamber with a glass bottom which fits upon the stage of 
the microscope, while the upper surface is a truncated cone of glass pro- 
jecting downward into the moist chamber. The lower surface of this is 
just 4 mm. in diameter on which is placed the drop of blood, care being 
taken that it only just covers the surface, hence is always of the same size. 

4 Loc. cit. 

5 Quoted from Ash, Arch of Int. Med., July, 1914, xiv, p. 8. 



THE BLOOD 



435 



The glass cone is then quickly fitted into the moist chamber. Through 
the side of this chamber projects a fine tube, through which, by means of a 
bulb, a gentle stream of air can be directed against the blood. With the 
low power of the microscope the cells are watched until they move in 
clumps. D E 





c 


1 -III 




^B^J 














c 


\ A 

C 






fe 



Fig. io6. — Coagulometer of Russell and Brodie as modified by Boggs. A, moist chamber; B, 
cone of glass the lower surface of which holds the drop of blood; C, side tube; D and E, cover- 
glass; at E, a pinhole. 

This method is the most accurate yet devised. The original apparatus 
of Russell and Brodie 6 has been modified recently by Boggs who uses a 
metal tube, an improved glass cone and dispenses with the water jacket 
(see Fig. 106). 

The corpuscles should be agitated as little as possible. They will at 
first move freely and independently of one another (see Fig. 107, A) and 
then in clumps on the periphery, B. As the process of coagulation con- 







FlG. 107. — Diagram to illustrate the movement of the cells during coagulation. 

tinues the masses of corpuscles will no longer move within the drop, but 
the drop changes its shape en masse, the corpuscles showing first an elastic 
concentric motion, C, and finally an elastic radial motion, D. That is, 
the masses of corpuscles will be moved toward the center by the current 
of air and will quickly spring back to their original position when the 
current of air ceases. This is taken as the end point since only then can 
fibrin be demonstrated if the disk be quickly removed and the drop be 
touched to a piece of filter paper. All clots should be confirmed in this 
6 Jour, of Phys., May 12, 1897. 



436 CLINICAL DIAGNOSIS 

way. Sometimes a " vicious circle " is set up in the drop, which clots 
everywhere but one point where the blood remains fluid. This is the result 
of too hard blowing. Such a drop should be discarded and another attempt 
made. 

Successive records at intervals of 5 to 10 minutes should not vary over 
30 to 45 seconds 

In estimating the coagulation time one must take into account not 
alone the method and the instrument, but also one's definition of the end- 
point. Using the Boggs instrument, Hinman and Sladen found it to vary 
from 3 to 8 minutes, averaging 5 minutes and 6 seconds. This is a longer 
time than some others have reported but they chose a late end-point. 
(Brodie and Russell reported 3% minutes; Murphy and Gould, 3 minutes, 
1 1 seconds; Pratt, 4 to 5 minutes.) Above 9 minutes certainly would mean 
delayed coagulation. 

HowelVs Method. 1 — Two or four cubic centimeters of blood are obtained 
directly from a vein by means of a sterilized syringe and expelled at once 
into tubes with a diameter of 21 mm. which had been cleaned carefully 
with a bichromate acid mixture (see page 15). The coagulation time is 
the period which elapses between the time the blood is obtained and that 
when the clot is firm enough to allow the tube to be inverted. Controls 
are always made using blood known to be normal. The needle should 
enter the vein at the first puncture since several attempts will allow some 
tissue juice to mix with the blood which will hasten clotting. A further 
development of Howell's work is the determination of the prothrombin 
time. 8 Using these-' methods Miss Pettibone 9 has made a most careful 
study of this subject which certainly will in the future have considera- 
ble clinical value. 

So many methods of such varying value have been used in studying 
disease conditions that the results are scarcely comparable. Most agree, 
however, that in hemorrhagic diatheses the coagulation time is immensely 
increased; to from 10 to 15 or more minutes in certain of the purpuras 
(due to a deficiency of platelets (Pettibone)) and to even 50 minutes in 
some cases of hemophilia (due to a deficiency in prothrombin) . In long- 
standing jaundice the coagulation time is increased (due to a calcium 
deficiency) and any operation should be delayed on such a patient until it 
has been decreased to about 5 minutes by proper medication. The coagula- 
tion time is diminished in venous stasis due to any cause, after repeated 
hemorrhages (in a recent case following postpuerperal hemorrhages for 
several days an ordinary blood count could not be made, so rapidly did the 
blood clot in the capillary of the mixing pipet), after transfusion, by hun- 
ger, by too long administration of calcium chloride and by carbon dioxide. 

7 Arch, of Int. Med., 1914, vol. xiii, p. 80. 

8 Arch. Int. Med., 1914, xiii, 76. 

9 Jour, of Lab. and Clin. Med., Feb., 191 8, III, p. 275. 



THE BLOOD 



437 



Bleeding Time. — The determination of the bleeding time should be made as a con- 
trol of the coagulation time since it will detect some cases in which operation might be 
serious because of hemorrhage and yet the coagulation time normal. 

Dukes' method is as follows: The lobe of the ear, cleansed as for obtaining the 
blood for the coagulation time, is pricked deeply, the time noted, and then each drop as 
it collects on the skin is picked off on filter paper (care being taken that this does not 
touch the skin) until the blood ceases the flow. This time seems independent of the 
depth and width of the drop provided only a capillary area is punctured. 

Fibrin Diagnosis. — In very thick smears of blood the fibrin strands may 
be seen to radiate through the specimen, usually from masses of platelets. 
These smears after standing for hours under a bell- jar are washed by a 
gentle stream of water which will remove the hemoglobin, 
the fibrin stained with fuchsin and the specimen dehy- 
drated and mounted. If examined fresh the specimen 
should be sealed with vaseline to prevent evaporation. 
Those diseases in which most fibrin is seen are pneumo- 
nia and acute articular rheumatism. In the former 
case this is suggested as a differential point against 
tuberculous pneumonia. 

The Viscosity of the Blood. — The best instru- 
ment in use is Hess's viscosimeter. On a base of opaque 
glass H (Fig. 1 08) are fastened 2 graduated glass tubes, 
A and B, which are connected at one end by a third 
tube, G, while this cross tube in turn connects, through 
a branch, with a rubber balloon, L. At their other end 
are 2 tubes, C and D, which are drawn to capillaries of 
very fine caliber, which widen again to the bore of A 
and B. The tube F, placed on H, and held there by 
the support N, is removable and can be replaced by any 
one of a number of similar tubes. By means of the stop- 
cock Q it is possible to establish or to interrupt the 
communication between B and the balloon L. The tubes A and B are 
bent to a right angle at their junction with G. Interposed between the 
rubber tube K and the balloon L is a glass tube which by an opening com- 
municates with the air. 

The method of making the determination is as follows: In the tube 
B-C-D is a column of distilled water the left meniscus of which is at 0. 
The tube F is filled by capillary action with blood. It is then placed end 
to end with D and by means of the suction produced by L the blood column 
is drawn up to 0. The cock Q is then opened and by the suction of the bulb 
the water and blood flow through the tubes A and B. As soon as the blood 
reaches 1 suction is discontinued and the readings are made. The body 
of water which has risen in B gives the relation of the viscosity of the blood 
in question to that of distilled water. The water and blood are now 
expelled by pressure on the bulb L, the cock is closed when the water reaches 




Or * 
Fig. 108. — 
Hiss' Viscosimeter. 



438 CLINICAL DIAGNOSIS 

O, F is removed and the tube DA is cleaned by drawing ammonia through 
it twice. 

If the blood is very viscid or coagulates rapidly it may be drawn to 
y 2 or %, and the values obtained multiplied by 2 or 4 respectively. 

Controls made with fluids of known viscosity are accurate to within 0.5%. 

Experiments by Hess showed that with a rise in temperature of i° C. 
the viscosity decreased 0.8%. Observations at temperatures of ordinary 
rooms show an error of about 4%, which is practically negligible. The 
only corrections of importance are those necessitated by great variations 
in temperature. 

The blood is obtained from the lobule of the ear, which has been previ- 
ously cleaned with alcohol. If the viscosity of the plasma is to be deter- 
mined the blood is drawn from the median basilic vein by venepuncture, 
coagulation being retarded by the addition of dry hirudin, and the 
blood sedimented. 

The viscosity of the blood in health is a variable factor. It is slightly 
greater in men (4.55) than in women (4.51); it depends on the number of 
red corpuscles, on the hemoglobin contents, on the gaseous richness, and, 
to a lesser degree, on the proteid, fat, and salt contents of the blood. And 
yet it varies directly with none of these factors. The viscosity of the normal 
plasma varies from 1.7 to 2.0, average 1.86. 

The results which Austrian 10 obtained are as follows : The viscosity of 
the blood and of the plasma are reduced in the anemias, both the primary 
and the secondary. In leukemia there is hypo viscosity of the blood and 
hyperviscosity of the plasma. The viscosity of both is increased in poly- 
cythemia. Hypo viscosity of the blood and hyperviscosity of the plasma are 
almost constant in cases of nephritis, the former because of the anemia and 
the latter to retained products of metabolism. Hypoviscosity occurs often, 
though not always, in cases with arterial hypertension. In cardiac diseases 
without edema no constant change is to be found, the coefficient apparently 
varying with the anemia and with the carbondioxide content of the blood. 
In cardiac cases with hydremia there is hypoviscosity of the plasma. In 
diabetes mellitus the viscosity of the blood and that of the plasma are 
increased. This may be the result of the hyperglycemia, of the lipemia and 
of the concentration of the blood due to the polyuria. In icterus both 
that of the blood and of the plasma are generally increased. In typhoid 
fever it varies with the anemia. It is increased by hydrotherapy and appar- 

TT-U 

ently is uninfluenced by diet. The -t-t quotient is more often decreased 

than increased. In pneumonia the viscosity is generally above normal. 

This may be due to cyanosis and to the retention of salt. Here, too, 

TTb 

quotient is low. In malarial fever the viscosity of the blood is usually 

10 Johns Hopkins Hosp. Bull., Jan., 191 1, vol. xxii, p. 9. 



THE BLOOD 



439 




normal or subnormal but is rarely above normal. That of the plasma is 
normal or increased if hemoglobinemia is present. In no disease can a 
pathognomonic alteration in the viscosity of the blood be demonstrated. 

The Estimation of Hemoglobin. — The estimation of hemoglobin should 
be the most satisfactory of the blood examinations but the use of faulty 
instruments has resulted in the accumulation of a vast amount of data of 
very little value. 

It is unfortunate that hemoglobinometers have not from the first been 
graduated to read in terms of grams per ioo c.c. of blood 
rather than per cent., for their makers have not agreed 
what quantity of hemoglobin should be called normal, 
nor would the same figure be normal for all ages. The 
blood of a normal child of about 10 years would read but 
80% with an instrument standardized to read 100% for 
a normal man of 30 years, etc. 

Another source of error is that many instruments 
are standardized against dilute water solutions of hemo- 
globin while hemoglobin in an albuminous fluids like the 
blood-plasma will give higher readings ; hence in reading 
the blood in extreme anemias we get misleading figures. 

There are at present but 2 instruments to be recom- 
mended, Miescher's hemoglobinometer and Sahli's hemo- 
meter, but so much of the past work has been done with 
other instruments that we will describe briefly a few of 
the more popular ones in order that the student may read 
the literature of medicine with better understanding. 

Miescher's Hemoglobinometer. — For years the 
Fleishl instrument was the best and this later was im- 
proved in some details by Miescher. Miescher's instru- 
ment is suitable for the laboratory and clinic only since 
it is expensive, bulky, requires a dark room, considerable 
time for each determination and considerable practice. 
The blood is diluted in a beautifully made melangeur 
(see Fig. 109), which allow dilutions of 1 : 200, 1 : 300 
or 1 : 400. These pipettes are particularly well marked, each small 
line on either side of the main lines indicatng Xoo of the length of the entire 
column thus saving the time necessary to bring the blood column exactly 
to one mark. 

A large drop of blood is aspirated to the point indicated for the desired 
dilution and then the pipette is filled with a 0.1% sodium carbonate solu- 
tion (A stock solution of 10% is diluted 100 times.) The dilution chosen 
should be such that the readings will be made near the middles of the color- 
prism. The pipette is handled exactly as for a blood-count (see page 456). 
One side of a cell (of which there are 2, one 15 mm. and the other 12 mm. 



w 



Fig. 109. — Mixing 
pipette of Miescher's 
hemoglobinometer. 



440 



CLINICAL DIAGNOSIS 



in depth) is filled with water. None should leak into the other half. The 
blood, well shaken is then blown into the other chamber of this cell. Both 
the water side and the blood side should have convex meniscuses. The 
cover-glass E is then slid in place pushing off the excess of fluid and leaving 
the chambers exactly full. The small cap, F, will now hold the cover-glass 
secure and also limit the field of vision. The cell is inserted in the receptacle 
on the stand, A, and the instrument placed in a screen which admits light, 
a yellow flame, whether from gas, oil, or candle, at one point only, where 
it will fall directly on the mirror and illumine both fields equally. Electric 
light, a gas-light with a mantle, or sunlight, cannot be used. The observer 
sits in a comfortable manner with the eyes about 25 cm. above the instru- 
ment and makes his observations with both eyes open rotating the milled 




Fig. 1 10. — Color-prism of the Fleischl-Miescher instruments. 

head C, which moves the color-prism, until that part of the prism (see 
Fig. no) which just matches the color of the blood-mixture is under the 
water-half of the cell. Since the retina is soon fatigued the eyes should 
be rested each 15 seconds. At least 5 different readings should be taken 
and the mean, not the average, used. The blood is then transferred with 
the melangeur to the shallow chamber and a similar series of readings is 
made for control. Since these cells have heights which are to each other 
as 5 is to 4 different parts of the color-prism will be used. The average of 
the readings with the lower cell multiplied by % should not differ from the 
average made with the higher by over 2%. 

Each instrument is accompanied by a scale which gives the number of 
milligrammes of hemoglobin per liter of diluted blood corresponding to the 
readings of that particular instrument. It is then easy, making due allow- 
ance for the dilution, to determine the number of grammes of hemoglobin 
in 100 c.c. of blood. If then, with due observance of the age curve, the 
worker wishes to express his answer as a percentage, he is at liberty so to do. 
This instrument is claimed to be correct within 0.2% of hemoglobin. 

The melangeur is cleaned, etc., just as is that of the blood-counter. 

FlcischVs Hemoglobinometer. — The pipette of the older Fleischl instrument was a 
small short cylindrical capillary tube (see Fig. in), which held from about 5 to 8 cmm. 



THE BLOOD 



441 



of blood. This was filled by just touching i end to a large drop of blood until the surface 
of the fluid at the ends is flat. Meanwhile, I side of the cell (see Fig. 112, H) of the instru- 
ment has been filled with water and a few drops of water placed in the other side. The 
pipette filled with blood is dropped into this latter side of the cell and emptied by rapidly 
agitating it in this water; and then washing any blood which may cling to the pipette 
back with a few drops of water from a medicine dropper. More water is then added and 
the whole well mixed by means of the handle of the pipette until this 
chamber of the cell is filled to the brim. The upper surface of fluid in 
the 2 halves of the cell should be just flat. They may be covered over 
with a suitable cover-glass. The instrument is read in a dark room, 
as is the Miescher. 

There are a few precautions to observe. The images of the 2 
halves of the cells should fall on the right and left halves of the re- 
tina, never on the upper and lower, since the lower half of the retina is 
not nearly as sensitive as is the upper ; the light should be at the side, 
never in front of the instrument ; as small a candle as possible should 
be used; if there is no screen handy, a tube of dark paper will suffice to 
cut out extraneous rays. The inconveniences of these machines are 
of ThV Fleischl in- that they use' a color prism which necessitates accurate standardiza- 
tion and a light of a constant color value. 
Gowers Instrument. — This little instrument (see Fig. 113) was for years the best 
the general practitioner had. It was cheap, easily portable, simple and when well made, 
fairly accurate. It consists of a color-tube. B, containing a fluid with the tint of a 
1% hemoglobin solution; a graduated test-tube, A, and a pipette C, which will measure 
20 cmm. of blood. The blood obtained in the pipette is diluted with water in the 



Fig. 



. — Pipette 




Fig. 112. — Miescher's modification of Fleischl's hemoglobinometer. A, stage; B, color-prism rack; 
C, Milled head; D, Cell; E, Cover-glass; F, Cap; G, Cell seen from above; H, Cell of Fleischl's Instrument. 

tube, A, until its tint matches that of B. The percentage is read directly on the gradu- 
ated scale from the height of the diluted blood. 

The color tubes contained gelatin stained with picrocarmine and so illumination of 
constant color value must be used. Each instrument had a tube to use in sunlight and 
another for gaslight. When not in use these tubes should be protected from sunlight. 

The Hemometer of Sahli (see Fig. 114). — For every-day clinical 
work Sahli 's hemometer is the best on the market. It is similar to Gowers' 
hemoglobinometer except that the color-tube contains a 1% solution of 



442 



CLINICAL DIAGNOSIS 



acid hematin, a pigment which is quite constant in composition and color 
value and the hemoglobin of the blood to be tested also is changed to acid 
hematin by hydrochloric acid. This instrument may be used in any light 
since the 2 tubes contain the same substance and would therefore be modi- 
fied by different lights equally. The blood, obtained in a graduated pipette 
holding 20 cm. (see Fig. 113, C), is blown into the graduated test-tube 
which previously had been filled up to the 10% point with a 0.1N HC1. 
(This may be made with sufficient accuracy by diluting 15 c.c. of the pure 
acid to 1 liter with distilled water. Sahli recommends that a little chloro- 
form be kept in this stock bottle.) The tube is thoroughly cleaned of the 





SS 




Fig. 113. — Gowers's hemoglobinometer. A, graduated Fig. 114- — Sahli's 

tube; B, color-tube; C, pipette. hemometer. 

blood by sucking up and blowing out the acid several times. The hydro- 
chloric acid will in a few minutes change the hemoglobin to acid hematin. 
It is then diluted with distilled water (mixing it well after each addition 
by covering the tube with the thumb and inverting it several times, then 
wiping back any fluid clinging to the skin by drawing the thumb across 
the mouth of the tube) until its tint corresponds to that of the standard 
color-tube. The instrument is provided with a very convenient little stand 
with a ground glass back which renders the reading easy and quite accurate. 



THE BLOOD 



443 



The color-tube certainly does deteriorate with age and so should be restand- 
ardized frequently. 

Dare's Hemoglobinometer. — This instrument (see Fig. 115), which has 
won a deserved popularity, compares a film of undiluted blood with a 
color-prism stained with golden purple. The pipette (see Fig. 116) consists 
of 2 plates of glass, 1 white, A, 1 clear, B, between which is a slit of known 
width. A rather large drop of blood is necessary and will at once by capil- 
larity fill the slit. The pipette is then at once slipped into the instrument 
(Fig. in, B), and the reading made before the blood can clot, using the 
light of a candle, E. The telescope tube, A, allows accurate focussing and 
also an advantageous magnification of the 2 color-fields. The prism is 
rotated by means of a small wheel, D, until the colors match and then the 





Fig. 115. — Dare's hemoglobinometer. A, telescope; B, pipette in 
place; C, case inclosing color-prism; D, milled head moving prism; 
E, candle; F, window admitting light to color-prism. 



Fig. 116. — Pipette of Dare's 
instrument. A, the white 
glass; B, clear glass disk. 



reading is made at the edge of the disk. The advantages of this instrument 
are that undiluted blood is used; that a determination takes but a few 
seconds; that leucocytes do not affect the reading as in the other instru- 
ments; and that it can be used in a light room. We have found that the 
readings of the same blood made by several persons with different instru- 
ments have compared very closely. 

The Tallqvist Scale. — This simple little book of blotting-paper with a scale of 
colors, exploited as a great boon to the general practitioner, has done great harm to the 
interests of accurate clinical work. The instrument can be carried easily in the pocket 
and a determination made in less than a minute. The colors of the scale vary by 10%, 
therefore any intermediate percentage must be estimated by the eye. We admit that 
an eye trained to use more accurate instruments will soon use this color-scale with some 
degree of accuracy but such a person would prefer to guess without its doubtful aid. 

The spot of blood is obtained by holding the edge of the blotting paper against a 



444 CLINICAL DIAGNOSIS 

large drop of blood and then at once blotting the paper by squeezing it between 2 pages 
of the book until the luster of the blood spot is lost. The reading is then made by reflected 
light before the drop becomes dry. 

In hospital work we find it a great advantage to have several varieties 
of hemoglobinometers in use and to insist that each student shall use them 
all. Only in this way can he learn to appreciate the strong and weak points 
of each. The man who uses but one soon places undue reliance upon its 
accuracy. Let him use 2 of different makes and find that they differ from 
5 to 20% and he will appreciate the difficulties involved. 

We have used a Miescher as standard and have required each clinical 
clerk first to standardize the instrument assigned him, which may be 
1 of 7 different types, against this and in future work to make the necessary 
correction. Later we have required the student to correct their instruments 
against several accurate blood counts on normal persons so that the reading 
of 5,000,000 red cells should be 100% (e.g., if the count is 5,200,000 and the 
reading 85, then 5,200,000: 5,000,000 : : 85 : x=ioo. Of course several 
such tests should be made.) 

These instruments very seldom read 100% for the blood of a normal person. We 
have careful records of 176 medical students, all normal men during the third decade 
of life. The Fleischl instrument used in 161 cases read from 65 to 110%; of these, 136 
varied from 80 to 100% and 52 from 90 to 95%; the mean was 92.5%. Of the 156 
records made with the Dare instrument, which varied from 65 to 110%, 105 varied from 
90 to 100% and the mean was 95%. Of the 150 records made with the Gowers', which 
varied from 70 to 120%, 81 stood between 90 and 100% and the mean was about 92%. 

The blood of 125 students was examined using the Miescher instrument. (Prac- 
tically all of these estimations were controlled at the same hour of the following day.) 
The results varied from 1 1 .4 to 17.6 gms. per 100 c.c. It was almost impossible to deter- 
mine the mean of these figures, so uniform was their distribution. Their average was 
about 14.5 gms. Considering this 100%, the blood of 38 students varied from 90 to 
100% and 21 from 100 to 105%. (Note how different the readings on the other instru- 
ments.) The range of variations with the Miescher seemed wide, yet they ran more 
parallel to the blood-counts than did the results with other instruments. 

Hemoglobin. — One hundred cubic centimeters of normal blood, it is 
usually stated, contain from 13 to 14 gms. of hemoglobin. Careful estima- 
tions of the hemoglobin at the various ages have shown that there is a defin- 
ite curve which runs quite parallel to that of the red blood-cells. 

Age Gms. per 100 cc. of Blood 

I to 4 days i9-3 2 9 to 21.160 

8 to 14 days 17.869 to 16.124 

8 to 20 weeks 15-362 to 12.928 

6 months to 5 years 10.971 to 11.373 

5 to 15 years 11-151 to 11.796 

15 to 25 years 13-034 to 13.870 

25 to 45 years 14-727 to 15.013 

45 to 60 years 12.484 to 13.150 

From this table of Leichtenstern (modified from Sahli) it is at once 
evident that the age curve must be considered in all blood- work, and that 



THE BLOOD 445 

it would be better were the hemoglobin estimations given in grams per 
ioo c.c. rather than in percentage, since there is no one figure which could 
be considered 100% for all ages 

By oligochromemia is meant a relative diminution in the amount of 
hemoglobin per unit volume of blood. It therefore is a relative and not 
an absolute value. 

THE EXAMINATION OF THE FRESH BLOOD 

The examination of the fresh blood in every possible case is a matter 
of routine which will take but about 3 minutes and may save a great deal 
of time. In the majority of cases not seen in the office we must rely on 
stained specimens, although the fresh blood would give more information, 
sometimes unexpected and of the highest value (see page 658), but more 
often hints which suggest to the worker along what further lines of examin- 
ation to proceed. It should also be a matter of routine to make a few 
dried specimens whenever one examines a fresh specimen or counts the 
blood. These need not be stained but can be filed away. Later, because 
of the further developments of the case, those old smears may prove of 
greatest interest. 

Technic. — To make fresh blood specimens, the slide and cover-glass 
must be perfectly clean (see page 463). The slide should be warmed by 
rubbing it rapidly with a cloth or by holding it for an instant near a flame, 
since the blood spreads much better on glass warmed to about body temper- 
ature than on cold glass. The skin is punctured, the first drop wiped off, 
and the second or a later one, when about 2 mm. in diameter, is picked up 
on the cover-glass held in the pinch forceps, care being taken that the cover - 
glass does not touch the skin, and this is then dropped onto the slide. The 
drop should be so small that when well spread the blood film hardly reaches 
the edge of the cover since the distribution of cells varies at different parts 
of the spread and so one should be able to examine the entire specimen. 
The blood should spread evenly. Under no condition should pressure be 
made on the cover-glass to aid the blood to spread since this will make 
a poor specimen only worse. Neither should the coverglass be pushed into 
a better position. The student should always be careful to drop the cover- 
glass on the convex side of the slide, and thus avoid a rocking specimen. 

Red Blood-cells. — In the well-made specimen the red corpuscles will 
all lie singly, flat on their sides, not overlapping, nor in rouleaux. If, as 
sometimes is the case, it is important to know whether the tendency to 
rouleaux formation is increased or diminished a larger drop of blood is used. 

The number of the red blood-cells may be estimated with a certain 
degree of accuracy by one who always uses approximately the same sized 
drop of blood. The shape of the red blood-cells is of considerableimportance. 
In the circulation they may, as is claimed, be cup-shaped, but in a well made 
specimen they flatten out on the glass as perfectly round or, where more 



446 CLINICAL DIAGNOSIS 

crowded, polygonal biconcave discs. In badly made specimens, and sooner 
and later in a good one especially along the edge, many of the cells are 
crenated, that is, are spherical and covered with small prickly points. If 
a cell has but one of these points and that is on the flat surface of the cor- 
puscle it may be mistaken for a small ring form of the malarial parasite. 
By crenation, however, is meant not alone this artefact of prickle formation 
but more especially a pathologic change in the contour of the corpuscle 
while in the circulation. Instead of being a round disc with a circular 
edge these cells have an uneven, shrunken margin. This is seen in the cells 
harboring the parasites of quartan and aestivo-autumnal malaria. 

The presence of poikilocytes is important in diagnosis. Poikilocytes are 
corpuscles which seem to lack that remarkable elasticity of shape which 
keeps them always round except when under direct pressure and so they 
are flabby and assume many odd shapes. Some are indented, some are 
ellipses, crescents or ovals while others have the well known sausage and 
battledore forms. 

Poikilocytes may be due, first, to technic. Pressure on the cover-glass 
will cause a certain number of the corpuscles to break up into small spherical 
masses and small elongated rods which resemble bacilli. The ease with 
which this occurs will depend to a great extent on the condition of the cor- 
puscles. If, because of disease, they are " weak," a slight injury will affect 
them more than normal corpuscles (Stengel). Any motion of the cover- 
glass after the cells have spread will distort them considerably. Second, 
heat will produce poikilocytes. If a specimen of fresh blood in a moist 
chamber be heated to from 50 to 54 C. the cells will present a most remark- 
able picture. The corpuscles lose their shape and show definite contractile 
movements. Some will elongate considerably and move around with a 
vermicular motion. We have known a whole hospital staff to study with 
astonishment the gyrations of these overheated red blood-cells, confident 
that some new parasites had been discovered. More commonly corpuscles 
when heated will bud, these buds become detached and swim in the serum 
as microcytes. Sooner or later in such a specimen nearly all of the poikilo- 
cytes will break up into fragments. Third, poikilocytes will increase 
the older a blood specimen gets. One assumse that red corpuscles are 
living cells, (although some believe that they die when they lose their 
nuclei before functioning as blood corpuscles) and that poikilocyte forma- 
tion under the microscope is evidence of death changes which appear much 
earlier than normal in the blood of some patients. These changes are best 
studied in well-sealed specimens on a warm stage, and resemble those of 
the over-heated specimen except that they are less in degree. But, finally, 
the poikilocytes which interest us most are cells which are misshapen in 
the circulation and so are seen in the quite fresh blood. A very few may 
be found in fresh normal blood, but many are present in that of any very 
severe anemia and especially in primary pernicious anemia of even mild 




d 



m 



H,J9ecKer 

fig 117.. — Fresh blood, a, cells with Maragliano's endoglobular degenerations; b, cell containing a 
navicular body, from a case of measles; c, the bacillus-like degeneration; d, a Maragliano degeneration 
in process of extrusion; e, a form of "hemoglobin degeneration" giving a dark area; /, like a; .?, like e\ 
h, a degeneration like e but almost free from cell; i, a pseudo "segmenting parasite"; k, an "ameboid" 
microcyte; I, estivo-autumnal hyaline malarial parasites; m, a full-grown estivo-autumnal parasite, 
and, n, a segmenter, both found in the peripheral blood; 0, same as l\ p, macrophage from a case of per- 
nicious malaria filled with malaria parasites. X 900. 



THE BLOOD 447 

grade. Of the many forms 2 were once supposed to be characteristic of 
this disease, those resembling a battledore and the elongated or sausage 
forms. Poikilocytes would seem to have ameboid motion; at any rate they 
changed their shape. This is best seen in the small ones. (Plate 1, 23-28; 
Fig. 117, &). The poikilocytes of anemia are probably immature forms 
rather than injured cells. 

The elasticity of red cells certainly varies; in lead poisoning it is said 
to be increased. The large pale cells in anemia on the other hand look flabby. 

The projection of the budding red corpuscles may have the color of the 
normal cells or be paler or darker. They are attached to the cell by a 
longer or shorter pedicle and often break loose. A former and mistaken 
theory of the origin of blood platelets was that they are free buds of red 
cells free of hemoglobin. 

The size of the red blood cells should be noted. Normally in the 
adult, these cells have a quite uniform diameter which averages 7. 5m- One 
always finds a few microcytes and some cells fragmented or injured by his 
technic. In the normal infant's blood there is much greater variation in 
size. In chlorosis these cells show quite a uniform diminution in size 
while in pernicious anemia cells of all sizes may be present yet the major- 
ity of them will be larger than normal. In secondary anemia they vary 
much in size and yet many will be normal and many others smaller than 
normal. In tertian malaria the infected cells are large, swollen and pale, 
while in quartan and sestivo-autumnal malaria the infected cells are small 
and shrunken. The average size of the red cells is said to be increased in 
jaundice, cholera, lead poisoning and leukemia; also in congenital heart 
disease and in cretinism. 

The color of the corpuscles is normally greenish-yellow. In the great 
majority of cases the variations in color are quantitative rather than quali- 
tative and due to differences in the amount of hemoglobin; that is, to the 
thickness of the corpuscles. The thickness of a corpuscle is easily estimated 
by the appearance of the cell at its center. While the biconcavity of a 
a normal cell can be made out only by accurate focussing that of a " light 
weight " corpuscle is very apparent while some cells are so thin that the 
hemogoblin cannot be seen at their center and they appear as narrow 
rings, the so-called "pessary forms." On the other hand, some corpuscles 
seem to lack a biconcavity while others, especially the microcytes, appear 
even biconvex. 

Some red corpuscles show a quantitative change of color, due apparently 
to some chemical change of the hemoglobin. For illustration, the corpuscles 
which contain quartan or estivo-autumnal parasites appear much darker 
than the other corpuscles and also have a greenish or " brassy " tone. A 
similar, although less marked, change in color is seen in some microcytes, 
in cells fragmented by mechanical injury and in red cells engulfed 
in phagocytes. 



448 CLINICAL DIAGNOSIS 

In some diseases the cells show a quite uniform change in color. In 
chlorosis, for instance, nearly all of the cells are paler and in pernicious 
anemia many will seem darker than normal. In other conditions it is the 
variation in color of the cells which is important, as in secondary anemia 
and in malaria. It is for this reason that the examination of the fresh 
blood is more valuable in diagnosis than that of stained specimens for 
in the latter much of color value is lost. 

Nucleated reds are often quite conspicuous in the fresh" specimen. An 
occasional normoblast is seen in normal blood but it is a pure anomaly. 

The partial degenerations of the red blood-cells are very important 
evidence of the intravascular health of the cells as well as dangerous sources 
of error in the diagnosis of malaria (Fig. 117). These are necrobiotic 
changes which appear sooner or later according to the intravascular condi- 
tion of the blood and the treatment it receives when or after the specimen 
is made. The areas in the cells showing these changes have received a 
variety of names, as vacuolization, pseudo- vacuolization, pseudo-nuclea- 
tion, etat cribriform, globular decolorization, but the name most commonly 
used is " Maragliano's endoglobular degenerations." These appear in 
normal blood as a rule in from 30 to 70 minutes after the specimen is made. 
Usually they develop near the center, but sometimes near the periphery, 
of the cell. One cell may contain one or several. The corpuscle appears 
thinner at one point and a vacuole-like area appears which seems free from 
hemoglobin. This area is usually round, although it may be elliptical, and 
increases in size until a mere rim of hemoglobin-containing protoplasm may 
be left. Although these spots resemble vacuoles they probably are areas 
of coagulative necrosis which may be extruded from the cell or remain 
visible when the rest of the cell goes to pieces. These areas certainly change 
their shape and their position within the cell, their rapid motions resembling 
those of malarial parasites. This is not true ameboid motion, however, 
but due probably to changes in the dying protoplasm around them. The 
rapidity with' which these degenerations appear in the specimen, other 
things being equal, will depend on the intravascular condition of the 
corpuscles. Maragliano and Castellino doubtless exaggerated their import- 
ance in diagnosis and prognosis, yet they are most conspicuous in severe 
cases of disease and are especially numerous in the primary anemias. These 
vacuole-like areas in the fresh blood and even more so in stained specimens 
may be mistaken for cell nuclei or for malarial parasites and explain many 
a mistaken diagnosis. Only the trained eye can distinguish them from the 
hyaline forms of the malarial organism, from which they differ in that they 
grow larger and more numerous the longer one searches for them. They 
occupy as a rule the center of the cell, are round or oval in shape, and, what 
is most important, they are too lens-like and enlarge or diminish in size 
as one changes the focus while a parasite would become less and less dis- 
tinct ; in general they are much easier to see than is a parasite ; their move- 



THE BLOOD 449 

ments may simulate those of an ameboid organism and their periphery 
may show the same wavy motion, but this is not true ameboid motion 
since they do not change their position by means of their change of shape. 
Some resemble beautiful " segmenters " (see Fig. 117, i). In Fig. 117 the 
attempt was made to show these differences (contrast a, /, and with /). 
In fixed specimens they show a granular structure and will take a basic stain. 

Their lack of constancy in size, the changes in their appearance on 
changing the focus and the absence of a distinct membrane and chromatin 
net-work of the nucleus should differentiate them from nucleated 
erythrocytes. 

Some special forms of endoglobular degeneration deserve particular 
notice; some which have a definite crescent shape are famous since twice 
described as the parasite of measles and more recently as that of spotted 
fever (see Fig. 117, b). 

Some rod-like areas resemble bacilli (c) . These may keep up a constant 
vibratory motion, moving practically through the whole substance of the 
cell and looking and behaving remarkably like a motile organization. 

Some present the appearance of a small dark cell on top of a larger 
and paler one, although focussing shows them to be in the same plane 
(see Fig. 117, <?, h, g, Ehrlich's hemoglobinemic degeneration). Another 
example of this degeneration is well illustrated in aestivo-autumnal 
malaria, cells whose hemoglobin is gathered in a mass around the parasite 
(see Plate V, 0), the rest of the cell colorless. This degeneration is best 
seen in pernicious anemia. 

Granules which would seem to be remnants of the nucleus and described 
by Vaughan n may be studied in fresh blood stained with Unna's poly- 
chrome methylene blue. To make the specimens the ball of the finger is 
well cleaned with alcohol and ether. On it is then placed a drop of the stain 
and the skin pricked through this drop in order that the cells may come in 
contact with the stain before they do with the air. A drop of the mixture 
of blood and stain is transferred to a slide and covered at once with a cover- 
glass. In a few minutes a few cells may be found containing violet granules 
which are coarse or fine, some in a line reaching across the cells and others 
connected by a filament. There is a remarkable constancy in their occur- 
rence. In normal adult blood they are found in from 0.5 to 1.8% of the 
red cells and in almost exactly the same percentage in a variety of diseases 
which have little influence on the blood. In the blood of the new-born, 
they are found in from 1 to 7% of the cells; in that of a fetus 2% inches long, 
in 24%. In the anemias they are more numerous, especially in primary 
pernicious anemia in which cases even 18.8% of the cells may contain 
such granules. Their number in general runs parallel to that of nucleated 
red cells. Vaughan gives as reasons for thinking that these granules 
are rem ains of the nucleus that they are not artefacts, they occur especially 

11 Jour, of Med. Research, 1903. 
29 



450 CLINICAL DIAGNOSIS 

in normal-looking cells, are situated in the position of the nucleus and are 
increased in conditions in which nucleated reds appear. He suggests that 
they may be a more delicate sign of anemia than are nucleated reds. 

Morris 12 has called attention to granules which are nearly always single 
and round, which are sharply circumscribed, eccentrically placed and which 
have the same staining reaction as the nucleus. These are almost certainly 
nuclear fragments. They are found in the blood of the human embryo, 
in the anemia of infancy and in those conditions of the adult in which other 
and clear evidences of blood degeneration are present, as pernicious anemia, 
secondary anemias, chronic myeloid leukemia, etc. 

Various poisons, potassium chlorate, pyrogallic acid, et al., often produce 
in the red cells vacuole-like areas or clumps, which are motile and which 
may break free from the cell, or which may be left free when the cell dis- 
integrates. Heinz and Bloch describe these as " areas of poisoned 
protoplasm." 

Leucocytes. — The presence of a leucocytosis and its character will 
often be suggested by the fresh blood examination, although such suspicions 
should always be confirmed by an actual count. To get a good idea of the 
leucocyte picture the fresh specimens should be carefully made since the 
distribution of the white cells varies somewhat in different parts of the 
specimens and so the entire specimen must be studied. 

The leucocytes in fresh specimens appear as colorless, nucleated, 
ameboid or immobile cells, which do not float in the current witn cne 
red corpuscles. 

In the fresh specimens the following leucocytes may be recognized. 
Small mononuclears which are cells about the size of a red corpuscle, some 
a little larger others smaller, with a nucleus relatively large, round as a 
rule although sometimes deeply notched and central in position. Their 
protoplasm is scanty. In some it is hardly seen while in others it presents 
a ragged edge around the nucleus and may appear somewhat granular. 
In certain conditions, e.g., in lymphatic leukemia, cells of this type are 
said to be ameboid and this is suggested by a study of these cells in tuber- 
culosis. Normally, these average 32.2% of the leucocytes. 

Large Mononuclears and Transitionals. Endothelial Leucocytes. — These, 
as seen in fresh normal blood, vary in size from that of a lymphocyte from 
which they can scarcely be differentiated in fresh specimens to others 
several times the size of a red blood-cell. The nuclei of these larger, very 
characteristic cells is sometimes round but more often oval in shape, eccen- 
tric in position and sometimes deeply notched (the " saddle-bag " or the 
■ ' wallet-shaped " nucleus). The protoplasm is very abundant and is clear. 
Although these cells appear non-ameboid yet it is interesting that in malaria 
they clearly are phagocytes. They average 7.2% of the total number. 

Ehrlich applied the term transitionals to those cells of the large mono- 

12 Arch, of Int. Med., March, 1909. 



THE BLOOD 451 

nuclear group which have deeply indented nucleus (see page 469). They 
would seem, however, to be merely the older forms of the endothelial 
leucocytes with which they are now counted. They are the largest of all 
blood cells. In their abundant protoplasm may be seen a few granules 
near the nucleus. 

Polymorphonuclear Finely Granular Cells. — The finely granular cells 
of Max Schultze average 58.5% of the total number of leucocytes. In the 
circulation they are from 10 to 15/* in diameter, but on the slide their 
apparent size is much larger and depends chiefly upon the extent to which 
these spherical cells are flattened out upon the glass. Their protoplasm is 
filled with fine dust-like granules. The nucleus has the shape either of a 
bent rod, a skein of fibers or of several masses of chromatin (hence the 
old name " polynuclear cells "). These when they leave the blood-vessels 
are the ordinary pus-cells, the greatest phagocytes of the body. 

The polymorphonuclear coarsely granular cells of Max Schultze, the 
eosinophiles are usually a trifle smaller than the preceding. Their nucleus 
is possibly less polymorphous but their protoplasm is filled with coarse, 
blackish, very refractive round or slightly oval granules of quite uniform 
size and shape, and about 1// in diameter. These are the most ameboid 
cells of the blood, and average 1.6% of the leucocyte count. 

The Mastzellen in the fresh specimen resemble the coarsely granular 
cells. While they cannot with certainty be recognized yet their granules 
vary much more in size and their nucleus is often trilobed. These cells 
average in normal blood 0.5//, of the total number. 

Pigmented leucocytes are best studied in the fresh or air-dried specimens. 
The pigment is sometimes from malarial parasites (melanin) and is very 
important in the diagnosis of malaria, or it is pigment picked up by the 
leucocytes in other conditions as in melanosarcoma. 

Hemosiderin pigment as ochre granules is seen, although rarely, in 
the leucocytes of cases in which there is rapid blood destruction. The 
iron of this pigment may be demonstrated by treating the smear first with 
2% potassium ferrocyanide and then with 0.5% hydrochloric acid. The 
specimen is mounted in glycerin. These granules will take on a blue color. 

Blood-dust or hemokonien granules is the name given by Muller to the 
very fine granules which dance actively between the red corpuscles in fresh 
normal as well as pathological blood. Finding them in large numbers in 
a case of Addison's disease he supposed that they bore some relation to 
that malady, but later found them present in all bloods, although in very 
varying amounts. They are small, round, colorless granules, which vary 
considerably in size, some even i/z in diameter resembling micrococci but 
the most much finer and dust-like. They are best seen by gas illumination, 
He found that they did not give the osmic acid test for fat, nor were they 
cleared by acetic acid, as albuminous granules would be. These granules 
were further studied by Stokes and Wegefarth 13 who decided that they 
13 Johns Hopkins Hosp. Bull., December, 1897. 



452 CLINICAL DIAGNOSIS 

were the free granules of leucocytes. Their reasons for this opinion were: 
that in man they resemble the leucocyte granules in size, being both coarse 
and fine, while the horse and rabbit which have peculiar granules in the 
leucocytes have similar blood-dust granules ; that they can be seen to escape 
from the leucocytes if certain reagents are added to the blood; and, lastly, 
the larger ones take an eosin stain. These granules are supposed to bear 
some relation to immunity. Doubtless all the so-called " spores " described 
in the blood are hemokonien granules. 

The fat globules of the body plasma appears in fresh blood specimens 
as exceedingly fine dust-like granules which would easily escape observa- 
tion. They form a perfect cloud in the plasma in cases of lipemia. 

The blood platelets in fresh specimens appear either singly, in clumps, 
or as masses of amorphous granules in the periphery of which are vacuole- 
like areas containing a watery fluid, the so-called " granular masses of 
Max Schultze. ' ' At this point we would remind the reader that all platelets 
in fresh blood specimens will at once stick to the glass and soon disintegrate 
and that a floating object in the plasma certainly is not a platelet however 
much it may resemble one (see page 525). 

The large macrophages can be studied only in fresh specimens since in 
stained preparations they appear as unformed masses of detritus. They 
may, in malaria, contain malarial parasites and red cells some of which 
may contain the parasites and in typhoid fever they enclose many red 
cells (see Fig. 117, ). 

In pregnancy placental cells (syncytium) swept off in the blood-current 
" are commonly found " in the mother's blood (Veit). 

The fibrin net-work often radiating from small masses of platelets seen 
in fresh blood preparations is of value in diagnosis. The amount of visible 
fibrin is very large in certain diseases, as pneumonia, acute articular rheu- 
matism, etc. 

Counting Red Corpuscles. — The blood-counting apparatus (see Fig. 
118), consists of a pipette in which the blood is diluted, a counting chamber 
by means of which a layer of the suspension of corpuscles of known depth 
and area is obtained and a special cover-glass to serve as the upper boundary 
of this layer. This apparatus should be carefully standardized and the 
necessary corrections always made for we have bought expensive pipettes 
from good dealers which had an error in calibration amounting to 40% and 
counting chambers with ruling definitely inaccurate. The pipette is a 
graduated capillary tube (Fig. 118) A, opening into a dilation, B, at the 
opposite pole of which is a second shorter glass tube, D, to which is attached 
a rubber tube with a mouth-piece. The pipette is so graduated that the 
capacity of the reservoir measured from the line marked 1 to that on the 
shorter tube marked 101, is exactly 100 times the capacity of the capillary 
tube from its point to the line marked 1 . As a rule the unit length of the 
long tube is divided into 10 sections but the only marks of importance are 



THE BLOOD 



453 



the i and the 0.5. We much prefer those pipettes which have on either 
side of the 0.5 and the 1 marks 2 smaller marks, each indicating the Moo 
length of the tube (see page 519), since then one need not bother to draw 
the blood just to the main line but using these short lines and calculating 
the correction can work more rapidly and therefore more accurately. The 
point of the long tube should not be 
too sharp since in the quick move- 
ments made it will be easily broken. 
In the bulb is a small ball, C, which 
aids much in mixing the blood and 
the diluting fluid. The pipette is 
cleaned by washing it out first with 
water, then with alcohol and then with 
ether Air is then sucked through, not 
blown, until the bulb is visibly clean 
and the glass ball rolls freely within it. 

Boggs u has improved the pipette by in- 
serting in the rubber tube a Wright's "throt- 
tle capillary." This consists of a capillary 
tube which has been heated in a very small 
flame and then quickly drawn out into a fine 
thread and an outer, protecting tube from 5 
to 7 mm. in diameter and of hour-glass shape. 
The large part of the capillary is marked 
with a file, so that it may be conveniently 
broken off after it has been cemented in the 
holder. The cementing is easily done by 
molding a little sealing-wax near the throt- 
tled end, passing the larger, free end of the 
capillary first through the holder, and, after 
warming gently at the constricted part, 
drawing the waxed end down into the nar- 
row waist of the tube. The wax softens, fills 
the neck of the hour-glass tube and on cool- 
ing leaves the capillary firmly cemented in 
place. Each end should be about 5 mm. 
shorter than the container. The larger end 

of the capillary is then broken off the point F . 1( f- 118.— Blood-counter (Thoma-Zeiss;. To the 

, . . _ . TTT1 right is the ordinary form of pipette for red cells; 

marked, Using a pair Of fine forceps. When the other is a leucocyte pipette with improved 

the pipette is washed this controlling de 
vice should be removed. 




leucocyte pipette with improved 
markings and point, D. The ruled counting cham- 
ber is shown on edge and face view. A, the slide; 
This controller B - the rin S; D > the ruled table and C, the "ditch;" 
... 1 r 1 1 -> E> trie cover-glass. 

makes it easy to draw the column of blood 

steadily and slowly to the point desired and also keeps the blood from falling from the 

pipette when the tip is transferred to the bottle of diluting fluid. 

Dr. James Wynn has further improved the pipette by providing the rubber tube 
with a double roller device which makes suction with the mouth unnecessary (Fig. 119). 

The apparatus consists of a hair pin (preferably a crimping pin the wire of which is 



14 Jour. A. M. A., January 5, 1907, vol. xlviii, p. 47, 



454 



CLINICAL DIAGNOSIS 



about o.i cm. in diameter) and 2 rubber cylinders, shaped with pen-knife from an ordin- 
ary Eberhardt Faber " Ruby " eraser and then smoothed with fine sandpaper. The 
shorter cylinder is approximately 0.8 — 1.25 cm.; the longer is of similar dimension in 
the center, but about 0.3 cm. longer at either end, these extremities being slightly greater 
than 0.8 cm. in diameter. This gives the longer cylinder a spool shape and enables its 
curved surface to exactly approximate the corresponding surface of the other cylinder. 
A hat pin is carefully thrust as near as possible through the axis of each cylinder 
and the 2 are worked onto the arms of the hair pin, reinsert- 
ing the pin shafts several times in order -to enlarge the axis 
passages until the cylinders rotate fairly easily. The cylinders 
are then pushed well up on their respective shafts and the 
curve of the pin is bent so that corresponding surfaces just 
touch. The pipette tubing is then slipped between the cyl- 
indrical rollers. In use the rollers are gripped gently between 
thumb and index finger, and rolled away from the attached 
end, compressing the rubber tube between them as they go. 
The vacuum thus created enables the operator to control 
with finger tip accuracy the rise of blood in the tube. When 
the blood mark has beenreachedthe tip of the tube is plunged 
in the diluting fluid and enough drawn in to empty the capill- 
ary of blood. The rubber tube is then released from between 
the rollers and using mouth suction the glass bulb almost 
filled. (This enables one to rotate the bulb while filling it etc.) . 
The tube is then gripped by the rollers as before and the di- 
luent graduation is reached promptly and exactly with no 
danger of overflow into the rubber tube. 

The worker will save much time if he pays 
careful attention to his pipette. It should be 
thoroughly clear before it is used, the capillary should 
contain no trace of clotted blood, the glass ball 
should roll freely in the bulb and the rubber tube 
should be very flexible, not cracked near the 
mouth-piece, and should contain no saliva. 

Of the diluting fluids in use Toisson's is the best. 
The formula of this is : 

Water (distilled), 

Glycerin (neutral), 

Sodium sulphate, 

Sodium chloride, 

Methyl violet, 




Fig. 119. — Wynn's roller de- 
vice for rubber tube of mix- 
ing pipette. 



160 C.C. 
30 C.C. 
8 gms. 
1 gm. 
0.025 gm., or just enough 
to give desired tint. 



Hayem's fluid is preferred by some : 
Distilled water, 200 c.c. 

Sodium chloride, 1 gm. 

Sodium sulphate, 5 gms. 

Mercuric chloride, 0.5 gm. 

Sodium chloride can be used in rather strong solution (3%). It is 
probable that the physiological 0.6% solution will lake a certain number 
of corpuscles. 



THE BLOOD 455 

These diluting fluids when used must be fresh and recently filtered since 
yeasts grow in them which lead to error. (We remember one case with 
normal leucocyte count in which a count of 110,000 cells was reported.) 

Some of the counting chambers consist of a heavy glass slide, (Fig. 1 18) A, 
on which is cemented a thick glass ring, B, the surface of which is carefully 
polished. This ring surrounds a circular table of glass, D, upon which is 
the ruled area and the height of which is just 0.1 mm. less than that of the 
surrounding ring. Between this ruled glass table and the inner edge of the 
ring is a small ditch or moat, C, to catch the drop of diluted blood which 
may run off from the table and to prevent this from running up between 
the ring and the cover-glass on the other side of the ditch. The later models 
(see Fig. 120) are more convenient since they have, instead of the thick 
glass ring, 2 parallel tables between which is a third just 0.1 mm. lower 
which is a narrow rectangle the ends of which reach nearer the sides of the 
slide than do those of the 2 higher 
tables. Those with open moat have 



■ r 



the great advantage that the cover- | ; : 

glass ma}-- be accurately adjusted 

and then the diluted blood allowed 

to run up beneath it by capillarity. ^ mam ' ™iri™afflr 

There is, therefore, no danger that I 

the chamber must be cleared up 

several times before Newton's 

band's appear. 

On the central glass table of the ' i 

older model and m the middle or the 

9 c/Ipqq tpblpQnft'hp IfltfprflTV* mlprl or FlG - 120. — Counting chambers with open moat 
2giaSStaDieSOimeiaT7GerarerUiea2I (Buerker). A, with single ruling; B, with double 

parallel lines, 0.05 mm. apart. Cros- rulmgi 

ing these at right angles is an exactly similar set of lines. The result of their 
intersection is a 1 mm. square, divided into 400 equal small squares (see 
Fig. 121). Through each fifth row of squares is ruled an extra line. This 
extra line is not a boundary but merely aids the observer to keep his posi- 
tion in the ruled area Indicated, not bounded, by these extra lines, the 
square millimeter is therefore divided into 16 units of 25 squares each. 
Other rulings are in use all of which agree in that one square millimeter is 
divided into many smaller and easily recognized units. 

When choosing a blood-counter these lines should be carefully studied 
since certain makers have put on the market very imperfectly ruled slides. 
They should first be examined dry to make sure that the lines are complete 
and then, when covered with a drop of water that their sharpness may be 
determined; for we have seen lines which appear very distinct on a dry 
slide practically disappear when covered by a drop of water. 

Before use this counting should be washed with water and carefully 
wiped with a soft cloth (a coarse one will blunt the edges of the lines making 



456 



CLINICAL DIAGNOSIS 



them indistinct) care being taken that no lint be left on the surface of the 
glass ring. Alcohol or ether should never be used for this purpose since 
they will dissolve the cement which fastens the center glass table. 

The cover-glass is a heavy one with planed surfaces made particularly 
for this use. Ordinary cover-glasses should never be used for their surface 
may be uneven, they are seldom flat and are so thin that the capillarity 
of the drop will bend them slightly. 

Diluting the Blood. — After the ear or finger has been pricked deeply 
(see page 428) so that the blood flows freely without the assistance of pres- 











1 
















1 
































J 


V | 


















c 




































\* 
































































1 












































































































j 


















t 


^ 
































1 


j 














1 
































i 


























































































































n 




































is 

































































































































































































































































































































































































Fig. 121. — The one square millimeter ruled area, much magnified, show- 
ing the units in common use. 

sure, a large drop is allowed to collect on the skin and is rapidly drawn 
into the pipette to the mark 0.5 or 1 according to the condition of the blood. 
Normal blood should be drawn only to the ppint 0.5, anemic blood to the 
point 1 . If drawn too far the column may be shaken down somewhat by 
tapping the point of the tube against a towel or rubbing it against the end 
of the finger, but unless there is very little correction to be made the instru- 
ment would better be cleaned up again and the whole work started anew. 
It is for this reason that we prefer the special marking mentioned on page 
452, and also that devices like those described on page 453 are of value. 
If the column is of the right length the tip of the pipette is rapidly cleaned , 
either on the finger or by wiping it on a towel and plunged into a bottle of 
the diluting fluid which is now drawn up into the pipette until the mixture 
reaches the line 10 1. While drawing in this fluid the tube is held vertically 



THE BLOOD 457 

and rotated between the finger and thumb in order that the diluting fluid 
may mix at once with the blood as it enters and that no bubble of air may 
cling to the inside of the bulb. When the fluid reaches the mark 101 (an 
error of i mm. in this case would mean a negligible error of only 0.03%) 
the pipette is withdrawn from the diluting fluid, its ends closed by the 
thumb and first finger and it is shaken vigorously in a direction at right 
angles to its long axis for at least 1 minute. Two or three drops are then 
blown out in order to empty out all the fluid which has not entered into the 
mixture. If the blood is not to be counted at once the pipette may be 
sealed for several days by stretching a wide rubber band over its ends. 
When the count is to be made the shaking is repeated as vigorously as 
before. 

To fill the counting chamber the pipette is well shaken, the capillary 
tube emptied by blowing out 2 or 3 drops and a small drop, the size of which 
can be learned only by practice, is blown out upon the ruled table and 
covered at once with the cover-glass. Theoretically the drop should be 
just large enough to cover the ruled glass table and yet none flow over 
into the ditch, but practically some may run into the ditch but not enough 
to run up onto the surrounding ring. The cover-glass should be put in 
position at once. The best way to do this is, we think, to grasp it by 
2 diagonal corners, to place a third corner against the slide with the edge 
of the glass ring as a fulcrum, and then allow it to rotate down onto the 
drop. In this way no air-bubble is enclosed. 

The next step is to determine whether any dust or dirt between the 
cover-glass and the glass ring is increasing the thickness of the layer of 
diluted blood. This is done by holding the slide almost on a level with the 
face and toward the window in such a position that light is totally reflected 
from the surface of the cover-glass. If the cover and slide are in good 
apposition a beautiful spectrum band of colors (Newton's bands) should 
appear over the surface of the glass ring. Should these colors not be 
seen the cover-glass may be touched by some instrument (but not by a 
pencil). This may bring out the color bands. If they remain when the 
pressure is relieved the specimen is satisfactory. If, however, the concentric 
Newton's rings appear around the point of pressure and disappear when the 
pressure is removed the slide should be cleared up and another trial made 
since the increased thickness of the layer of blood will lead to great error 
in the final result. 

This test of good technic, the phenomenon of light interference, is to 
many a great bugbear but the fault usually lies in the counting chamber 
itself. We have bought slides on which the bands could almost never be 
obtained and others using which we seldom failed to get them. In the case 
of the newer instruments we get the cover in position first and then intro- 
duce the blood. While handling the glass slide it should be kept as nearly 
horizontal as possible since a slight tilting may allow the cover-glass to 



458 CLINICAL DIAGNOSIS 

slide off. The counting chamber should now be allowed to rest for from 
3 to 5 minutes in order that the corpuscles may settle onto the surface of 
the glass and therefore be counted more easily since all are in one plane. 

It will be seen that at certain points of this technic the movements must 
be very rapid. It is no exaggeration to say that greater mistakes are 
sometimes made by too careful than too quick work. It is of great import- 
ance that the pipette capillary be filled quickly, that the dilution be made 
rapidly (otherwise one finds groups of corpuscles not broken up by the 
shaking), that no time be lost between the final shaking and the blowing 
out of the drop into the ruled area and, in the older chambers, that the 
cover-glass be at once placed in position. 

The student who uses a chamber with closed moat will soon learn that 
it takes less time to clean up his counting chamber or his pipette and begin 
anew than to count a lot of extra fields with the hope of correcting some 
error which he is conscious to have made. One saves time by working 
with 2 counting chambers since the cells on one can settle while the other 
is being counted. 

It is also of great importance to examine the ruled area carefully with 
the low power before beginning the count in order to be sure that the cells 
are fairly evenly distributed over the table. If this is not the case the slide 
should be cleaned up and another preparation made. 

In counting the cells a medium high dry power of the microscope should 
be used by beginners, later the lower powers. A mechanical stage is often 
useful and yet it is better to train the fingers to move the counting chamber. 

The unit of the ruled surface (see Fig. 1 2 1 ) to use is a matter of individual 
preference. Cabot recommends 1 of 36 small squares, D; that is, a unit 
the 4 sides of which are rows of squares through each of which passes 1 of 
the extra lines. Simon prefers a unit of the 16 squares, B, through none of 
which the extra lines pass. Sahli recommends a unit of 4 squares, A. We 
count the 4 corner units of 25 small squares each of 1 specimen and then 
clean up the chamber, shake the pipette well, blow out several drops, fill 
the counting chamber again and count these same units. That is, we count 
8 units, or 200 small squares or % of a square millimeter. This is the least 
that a beginner should count. Later he may count the 2 diagonally oppo- 
site corner units of 2 preparations, or % of a square millimeter. When 
counting, those cells which touch even with their edge the upper and the 
left-hand lines are included in the unit, while those cells which touch in 
any way the right-hand or the lower boundary lines even though the cell 
lies entirely inside the square are to be disregarded. Since one counts 
downward and to the right there is less danger of counting the same cell 
twice if this rule be followed. The beginner should not try to avoid leuco- 
cytes but rather count them as red blood-cells. If in normal bloods all 
were counted as red cells the error would be but 0.09% which is of course 
negligible, although a high leucocytosis in a case of anemia would introduce 



THE BLOOD 459 

considerable of an error. In counting leukemic blood it is best first to count 
all leucocytes with the reds and then to count the leucocytes alone (see 
page 461). The difference will be the red cell count. Many of the red 
corpuscles will appear distorted and in some anemias the many very small 
cells are easily overlooked which may explain the very high color index in 
certain cases of pernicious anaemia. 

If the students counts 8 unit quares of 2 5 small squares each, then the 
sum of the cells counted multiplied by 2 will be the number of cells in a 
layer of diluted blood 1 mm. square and Xo mm. deep. If the blood was 
normal this would be about 2250. This multiplied by 10 will give the num- 
ber of cells in a cubic millimeter of the diluted blood, and this multiplied 
by 200 (providing the blood was drawn to the 0.5 point) the number of 
cells in a cubic millimeter of undiluted blood. This is the desired figure. 
In case, however, any other unit was used the calculation would differ 
accordingly. 

Our students are taught that if the extremes of the 8 counts, each the 
number of cells in a unit of 25 small squares, differ by over 25 cells they 
must repeat the entire count using a new blood mixture. If their technic 
is good it will be easy to fulfill these requirements but if a mistake has been 
made it is easier to clean up and begin over than to try, by counting more 
units, to offset an error due to poor distribution. 

We require third-year medical students to count the blood of 1 person 
daily at the same hour until the difference between the counts of 2 succes- 
sive days is not over 200,000 cells and the difference between the highest 
and lowest counts of the 8 units for each day not over 25 cells. That is, 
we allow an error of 4%, which is enough to include any physiological 
variations in the count and the error due to counting, which should not 
be over 2%. Some students attain this accuracy quickly. Some students, 
however, repeat this daily counting for from 20 to 30 or even more days 
before their work was satisfactory to themselves or to us. By this time 
they certainly have learned wherein lies the error in their technic. It is of 
interest that the most careful ones sometimes make the great errors, since 
they take too much time where speed is essential. Only those who have 
tested their own accuracy know how inaccurate they can be. One trained 
in the above manner is seldom guilty of reporting " rises or falls " of 
100,000 cells, nor would he ever report a count of e.g. 4,750,600. The student 
who knows that he can comply with this rule has a justifiable confidence in 
his technic. Blood-counting requires considerable practice. Even good 
workers after a vacation of a few weeks find that it is necessary to practice 
a little before they are ready again for accurate work. 

After the count is finished the slide should be washed with water only 
and dried with a soft rag, and the pipette is rinsed first with water, then 
with alcohol and then twice with ether. Air is then sucked through it 
until the glass ball rolls easily. If alcohol is drawn in before the blood is 



460 



CLINICAL DIAGNOSIS 



entirely removed an albumin precipitate will form. To remove this the 
pipette is filled with a pepsin-hydrochloric-acid mixture and left in the 
thermostat overnight. In case a clot obstructs the bore of the pipette it 
may be dislodged with a horse-hair. A fine wire should not be used for 
this will easiLy crack off the end of the tube. 

Trained workers have considered an error of 2% unavoidable and some 
are satisfied with 1 of 3%, which would mean that 2 men counting with 
equal accuracy the same normal blood at the same time might get results 
which differ by about 150,000 cells. We know no better way to stimulate 
students to attain good technic than by requiring a certain number of them, 
the more the better, each with a separate instrument to count independently 
the same blood. We have seen this result in considerable extra practice. 

The Hematocrit. — The hematocrit promised at first to save much of 
the time it takes to -count blood since by it we can determine the volume 




Fig. 122. — -Arm of hematocrit. 

of the red blood-cells, that is, the volume of the hemoglobin-containing 
protoplasm. This instrument is a centrifuge capable of very high speed. 
Each arm of the centrifuge (see fig. 122) holds a small glass tube of rather 
large bore calibrated with 100 divisions. One of these tubes is inserted 
in a rubber tube with a mouth-piece and the blood drawn in until the tube 
is even more than full. This requires a very large drop. The finger, 
covered with vaseline, is then placed over the free end and then the rubber 
tube removed. This glass tube is fastened in one arm of the centrifuge 
and -in the. other is placed an empty tube to balance the machine. The 
machine is then revolved at as high a speed as possible until the column of 
centrifugalized corpuscles does not decrease. Each division of the tube 
corresponds approximately to 100,000 cells. As a means of counting 
normal blood this method is fairly accurate but it is the abnormal bloods 
which it is important to examine and in these the variations in the size 
of the corpuscles introduces too great an error to be overlooked. But the 
instrument has its uses. By means of it we may determine the volume 
index of the red cells, the numerator of which is determined with this instru- 
ment (the count is the denominator). Capps 15 certainly has published 
some interesting results. By means of it we also may examine the plasma 
for bilirubinemia and for lipemia. 

15 Jour, of Med. Research, 1903, vol. vi. 



THE BLOOD 461 

Leucocyte Counting. — The leucocytes may be counted in the same prep- 
arations with the red cells, especially if Toisson's fluid was the diluent 
used. That is, after counting the red blood-cells one counts the leucocytes 
over the entire square millimeter. The trained eye will pick the most of 
them out, more because of the difference in their refractivity than from their 
stain, since they appear brighter when the focus is slightly raised. On the 
Thoma-ruled slide the leucocytes on the entire millimeter field of 8 separate 
drops should be counted. This requires considerable time and the number 
of cells counted is much too small, yet a fairly approximate result is ob- 
tained. It is much better to lake all the red cells by using as diluent i % 
acetic acid and to use the same pipette as for the red count but to draw the 
blood to the i line (giving a i : ioo dilution) or to use special pipettes 
which will give a dilution of i : 10 or i : 40 (see Fig. 118). The fluid is 
made up by mixing 1 c.c. of glacial acetic acid and 99 c.c. of distilled water. 
The 0.3% solution mentioned in several text-books is hardly strong enough 
since the red blood-cells will not be entirely laked and the groups of shadows 
left are confusing. This mixture should be made up fresh each day, for 
yeast-cells which resemble mononuclear leucocytes will grow in the dilute 
acid. It requires considerable practice to use these big pipettes. Their 
bore is so large that the blood easily drips out; it is difficult to wash the 
blood entirely into the bulb by means of the acetic acid; and in shaking 
it is easy to shake the leucocytes down into the fluid filling the tube. To 
reduce these errors as much as possible the pipette should be held almost 
horizontal while the blood and then the acetic acid are drawn into the tube. 
For this reason the bottle holding the acetic acid should have a wide mouth. 
The acid should be sucked in rapidly that the stream may wash the tube 
well. The pipette, with its ends firmly closed, is then shaken in all direc- 
tions except in that of its long axis. In this case also the specimen should 
be first inspected with the low power to make sure that the distribution of 
cells is even. 

If the counting slide has the Thoma ruling and but 1 sq. mm. area 
ruled and if the dilution is 1 : 100, at least 8 different slides should be pre- 
pared and counted; 5, if the dilution is 1 : 40. If however the ruling gives 
9 sq. mm. for the count (Fig. 123) then 3 specimens should be counted. 
At least 100 lenucocytes should actually be counted in each blood examina- 
tion made and more if possible. If the acetic acid is of proper strength, 
is fresh and if the pipette is clean all cells seen maybe counted as leucocytes. 
If the total number of cells counted be divided by the number of 1 sq. mm. 
units examined, this quotient multiplied by 10 and this by the dilution, 
the product will give the number of leucocytes in 1 cmm. of undiluted 
blood. If nucleated reds are present in the specimen their nuclei will be 
counted in the acetic acid as small mononuclears. It will, therefore, be neces- 
sary to determine their number relative to the number of leucocytes by the 
differential count of a stained specimen and then make the proper correc- 



462 



CLINICAL DIAGNOSIS 



tion of the leucocyte count. The hour the blood was taken for a leucocyte 
count should always be stated, the temperature of the patient at that time 
and whether or not the patient had partaken of a proteid meal. 

The student should be required to test his own accuracy. The error 
in leucocyte counting with standardized instruments is usually at least 5%. 
If a large number of leucocytes is counted it may be reduced to about 




Fig. 123. — A, Zappert ruling; B, Turk's ruling. 

3-5%- We are sure that the error made by the busy ward man is nearer 
20% and yet we hear, for instance, of " a rise of leucocytes from 10,000 
to 11,000 per c.mm.," etc. A careful man will by repeated controls make 
sure that the error of his technic is not over 5%. This can be done by 
filling several pipettes at the same time and counting them separately; 
or better, by inviting another in whose work he has confidence to make 
a series of parallel counts with him. In control 
work the blood should be taken at the same time 
and from the same incision, for one can obtain 
curious results if he takes the blood from (lif- 
erent parts of the body, especially if he uses the 
ear on which the patient has been lying or the hand 
which has been in a hanging position. 

Blood Smears. — To get satisfactory stained spe- 
cimens one must first get thin, well spread smears. The best method is 
that Ehrlich recommended. Two cover-glasses, % of an inch square and 
of the thinnest glass, are thoroughly cleaned in alcohol and ether (see page 
428) and then dried. One cover-glass is held along one entire edge by the 
crossed-bladed forceps. The other cover-glass is placed in a convenient 
position to be quickly picked up with the pinch forceps. A small drop 
of blood about the size of a small bead (about 1.5 mm. in diameter) is 




Fig 

from Osier) 



Platelets (copied 
a, platelets in ir- 
regular shapes; b, with clear 
areas; "c," Schultze's granu- 
lar mass. 



a 




- d 



f 




Fig. 124. — Nucleated reds from the blood of a fetus 15 cm. long, a, mature nucleated red; b, inter- 
mediate form and rosette; c, mature red, nucleus fragmented; d, free nucleus of a mature red; e, mature 
red, polychromatophilie cell;/, polychromatophilic megaloblast. 



THE BLOOD 



463 



picked up on the last mentioned cover-glass which is then at once dropped 
onto the other cover-glass. If the covers have been properly cleaned the 
blood will spread rapidly without the assistance of any pressure other 
than the weight of the upper glass. Just as the film is about to stop 
spreading, but before it has stopped the 2 covers are pulled apart in the 
direction of their planes by a steady but quick motion (see Fig. 105) 
which requires a little practice. Beginners may find it easier to hold the 
free cover-glass in the fingers but the moisture from the skin injures the 
specimens to a slight degree. With the 2 pairs of forceps one can make 
100 or more smears in less than 15 minutes. As soon as the covers are 
drawn apart they are allowed to dry in the air, not over a flame. They 
then remain spread out on a sheet of paper, blood side up, for from 15 to 30 
minutes to become perfectly dry, but must be guarded against flies which 




Pig 126. — Showing operator saturating blood plasma with carbon dioxide. 

suck up the hemoglobin, making large holes in the specimens. For some 
stains the smear is not allowed to dry but is dropped at once into the 
fixing fluid. Dry smears should be guarded from dust and moisture. 

Some prefer to use slides instead of cover-glasses. A large drop of blood 
is picked up on a slide and is spread at once by the edge of another slide, 
the ground edge of a glass spreader, the edge of a strip of paper or by 
drawing a needle flat across the slide. The use of slides has two advantages 
— no cover-glasses need be used, and a much larger blood surface is 
obtained for study. On the other hand only a few smears can be made at 
a time; more blood must be used; and the slides are bulky. But the most 
important objection is that the spreads cannot be as even. The leucocytes 
do not spread as do the red cells, but stick to the glass, some forms more 
quickly than others. In the areas too thick to study one may be sure there 
will be too many white cells relative to the red cells and relatively too many 
of some forms of leucocytes. Several smears have been sent us as illustra- 
tions of extreme leucopenia. In one case it was claimed that not a single 



464 



CLINICAL DIAGNOSIS 



leucocyte could be found, but further study showed many in the thick 
areas. Masses of leucocytes may be found on the areas first covered by 
the drop and give there the picture of an extreme leucocytosis. Even when 
2 cover-glasses are used the picture is not just the same on both. One 

cannot assume that the percentage relations 
seen in the thin areas is true of the thicker 
areas. We feel that one reason why differential 
counting has yielded such meager clinical re- 
sults is the use of slides and the consequent 
inaccuracies of the results. Even with the 
most careful technic many of the most in- 
teresting cells are usually ruined. We refer to 
the macrophages and to the very large mono- 
nuclear cells which can be found in fresh speci- 
mens and in sections of drops of blood hardened 
en masse. 

Air dried smears may be kept for some time 

if Ehrlich's stain is to be used but for the 

methylene blue-eosin mixtures now in vogue 

it is much netter to stain the smears at once 

**| P)l^ 1 I even before they are quite air-dry. 

Fixing Methods. — The fixing methods 
used will depend upon the stain to be em- 
ployed. Among the various methods are : 

Methyl alcohol is now the most popular fix- 
ing agent, since it can be used as the solvent 
of many stains and so allows fixing and stain- 
ing to be simultaneous. With other stains 
ethyl alcohol is used. 

Heat. — This method, the most difficult of all 
to use well, is the only one which gives satisfac- 
tory results with Ehrlich's triple stain. This 
is best done on a large triangle of unpolished 
copper plate, with a gas burner under the 
point. This is allowed to heat until at a constant 
temperature and then the boiling point is de- 
termined with drops of water. The cover-glass is 
placed on the copper plate with its outer 
fig. 127.— co 2 apparatus. margin (the margin farthest from the flame) 
% of an inch, blood side up, ins'de of the boiling point. Here the 
temperature is from no° to 115 C. This point can be determined more 
accurately by dropping toluol or xylol on to the plate since this :s their 
boiling point. The smears should be left at this temperature for an hour 
and a half to 2 hours. If the blood is heated on the day when the specimen 




THE BLOOD 



465 



is made 2 hours of heating will be hardly long enough. Specimens a week 
old generally require an hour and a half of heating and still older specimens 
less than an hour. The duration of heating depends also on the patient's 
disease. Normal blood requires the longest heating. Blood from a patient 
with splenomyelogenous leukemia is usually ruined if left on the bar more 




Fig. 128. — Apparatus for removing furnes in connection with nitrogen determinations. 

than 1 hour and to heat the blood of a patient with pernicious anemia a 
few minutes too long will often spoil the specimens. We place 4 smears 
on the bar at the point described above and remove the first in 1 hour and 
the others later at intervals of about 20 minutes. One of these is almost 
certain to stain satisfactorily. 

Others place the smears, blood side up, for from half a minute to 2 
minutes on the spheroidal point on the bar; that is, at the point where 
the drop of water just rolls off without boiling. Here the temperature is 
from 140 to 150 C. 
30 



466 CLINICAL DIAGNOSIS 

Success with Ehrlich's stain will depend in large measure on the heating; 
an over-stained smear is underheated and vice versa. The red cells are 
the best index of success. They must take an orange color with no fuchsin 
tint and yet not a lemon-yellow tint. 






:,.<; 'i-~-.ZMV' ' 




PlG. 129.- — Apparatus for Blood Cultures. 

Stains. — The stains were classified by Ehrlich as acid, basic, and neu- 
tral, not according to their chemical reaction but according to the nucleus 
of the stain which serves as the dye. 

The classical illustration of this is the following: ammonium picrate 
is an acid stain since it is the picric acid and not the ammonia which is the 
dye; rosanilin acetate is a basic stain, since it is the basic element which 
is efficient, not the acetic acid. Rosanilin picrate would be a good illustra- 
tion of a neutral stain since both the basic and the acid nuclei would stain. 
As a matter of fact the neutral stains are all mixtures of 1 or more stains 



THE BLOOD 



467 



A 



and it is very hard to state just how the compound arises and what it is. 

Among the basic stains may be mentioned methyl green, methylene 
blue, fuchsin, methyl violet, Bismarck brown, and safrranin. 

Among the acid stains, eosin, aurantia, the salts of picric acid, indulin, 
acid fuchsin, orange G, and a long list of others. 

Neutral stains arise in mixtures of the above. For in- a b 

stance, mixtures of fuchsin and methyl green; of methylene 
blue and eosin. 

(i) Hematoxylin Eosin. — This stain is not used nearly 
enough, since it, best of all, brings out the nuclei of the 
blood-cells. 

Mayer's Solution. — This stain contains hematoxylin 
i gm., alcohol ioo c.c, alum 50 gms. added while cool 
and then 1000 c.c. of boiling distilled water. A few crystals 
of thymol are then added and the whole cooled and filtered. 
It must be kept in the dark. One must determine by experi- 
ments how long to stain the specimens, after which they are 
washed rapidly in water. The nuclei alone will take the 
color. Eosin, 0.5% aqueous solution, may then be added 
until the red blood-cells are just rose red. The specimen 
is then washed in water, dried and mounted. The proto- 
plasm and the nuclei are beautifully stained but not all the 
granules. 

The polychrome methylene blue-eosin stains are at present 
practically the only ones in use. They are the only stains 
which bring out the chromatin of the malaria parasite; 
they stain the nuclei of the leucocytes very well and also 
the Mastzell granulation. The eosinophile granules are 
well stained but not so the neutrophile although perhaps 
as well as is necessary. If, however, one is studying granu- 
lations especially he will not use this stain alone, nor, 
indeed, any stain containing methylene blue, because it 
is so tricky. At least 16 different methods 16 of making this stain have been 
reported, all of them modifications of the original Romanowski's stain. 

Hastings' Mixture. 11 — The dry stains necessary are eosin, soluble in 
water, yellow (Griibler) ; and methylene blue (Ehrlich's rectif.) (Griibler). 

Solution A = eosin 1% aqueous. 

Solution B = alkaline methylene blue 1% aqueous. 

Solution C = methylene blue 1% aqueous. 

Solution A will keep, but solutions B and C must be made fresh. 

To prepare B add to a warm 1% solution of dry powdered sodium 
carbonate 1% of methylene blue powder and heat over a water-bath for 

16 Baumgarten, American Med., 1904, vol. vii, p. 14. 

17 See, also Wright's method, Jour. Med. Research, 1902, vol. ii. ,p. 139. 



! 



Fig. 130. — Tubes 
filled with clotting 
and clotted blood. 
A, blood is clotting 
spontaneously, the 
clot now retracting 
from the sides. B, 
clot in centrifugal- 
ized tube. 



468 CLINICAL DIAGNOSIS 

15 minutes. Add 30 c.c. of water for each 100 c.c. of original fluid, and heat 
again for 15 minutes. The solution is then decanted from the residue and 
divided into 2 equal parts. The 1 part is made faintly acid with 12.5% 
acetic acid, (this is best determined by placing a drop on blue litmus paper 
and taking as the end reaction the point at which the margin of the drop 
after absorption in the paper shows a faint pink) and then mixed with the 
remaining unneutralized portion. 

To make up the stain mix distilled water 1000 c.c, solution A 100 c.c, 
solution B 200 c.c. and solution C 70 to 80 c.c. In adding solution C, pour 
in 70 c.c. at once, stir well and if no precipitate is present add more, a 
cubic centimeter at a time, until one just appears. The stain is then allowed 
to stand for half an hour and then filtered through 1 filter. Forced filtration 
is usually necessary. The dry residue is removed from the paper and 
reduced to a powder in which form it may be kept. Seven to nine-tenths of 
a gram of dry stain is usually obtained. If more than 0.9 gm. the resulting 
stain is useless. The staining solution is made by dissolving 0.3 gm. of 
the dry stain in 100 c.c. of Merck's pure methyl alcohol. To do this the 
stain must be rubbed up with the alcohol in a mortar and pestle since it 
is with difficulty soluble. 

Usually the blood smear is covered with 2 drops of the stain for 1 minute, 
which will fix the specimen, and then 4 drops of distilled water are added 
and the dilute stain left on the smear for 4 minutes. For uniformity, a 
dropper should be used. These figures are merely examples. For each 
new lot of stain one must determine the relative proportions of stain and 
water to' be used in staining and the relative lengths of time to let the 
undiluted and the diluted stain act. 

Wilson's Stain. — One makes ai% solution of met hylene-blue in an 0.5% 
aqueous solution of sodium carbonate and adds at least 0.5% of freshly 
precipitated silver oxide. 18 The methylene-blue solution is boiled and at 
the end of 20 minutes % of it removed. After 20 minutes more boiling % 
of the liquid is removed and the remainder boiled for 20 minutes. These 
3 portions of fluid are combined and the mixture made equal to the original 
volume with distilled water, discarding the precipitate which sticks to the 
bottom of the evaporating dish. The methylene-blue solution is mixed 
with an equal volume of a 0.5% aqueous solution of eosin and allowed to 
stand for 1 hour. The precipitate is collected ona" hard " filter paper, 
washed with distilled water or preferably with 0.85% sodium chloride, 
dried and preserved in a dark glass bottle. 

To prepare the staining mixture 400 mgms. of the powdered stain are 
dissolved in 100 c.c. of absolute methyl alcohol. Since the powder is only 

18 To prepare the silver oxide, dissolve 2 gms. of AgN0 3 in 15 c.c. of distilled water 
and add about 260 c.c. of milk of lime. Shake well and set aside. Decant the super- 
natant fluid, collect the precipitate on a filter, wash it out with about 20 c. c. of dis- 
tilled water, dry at a temperature not exceeding ioo° c. and preserve it in a tightly- 
stoppered dark bottle. 



THE BLOOD '469 

slightly soluble the solution is facilitated by rubbing it up with the alcohol 
in a mortar. The stain should be kept in a tightly-stoppered dark glass 
bottle. 

The cheaper grades of methylene blue are said to make satisfactory stains. 

If the stain used is one in which the cover-glasses are to be completely immersed 
much time may be saved by using the holder which Pepper has invented. This allows 
55 slips 19 to be stained simultaneously in 35 c.c. of the staining fluid. 

For basophilic granules the methylene blue stains, carbol-thionin, or 
dahlia may be used. 

A good carbol-thionin mixture is: thionin 0.3 gm., absolute alcohol 
10 c.c. and carbolic acid, 1%, 100 c.c. The smear is stained in this for 2 
minutes, washed in water and dried. 

. A Specific Stain for Endothelial Leucocytes. — Several stains supposed 
to differentiate between lymphocytes and lymphoid cells or lymphocytes 
and endothelial leucocytes have been published, among them one proposed 
by Mcjunkin and Charlton 20 which brings out a granulation which they 
consider characteristic of the endothelial leucocytes. 

To prepare the solution one adds 0.2 gm. of alphanaphthol (Merck 
reagent), 0.015 gm. of methyl violet 5B (Grubler) and 0.2 c.c. of hydrogen 
peroxide to 100 c.c. of warm 80% alcohol (made from absolute alcohol). 
The hydrogen peroxide used should contain approximately 3% by weight 
of the gas as determined by titration with decinormal potassium 
permanganate. 

The blood film on a 22 mm. square cover-glass is covered for )i minute 
with 5 drops of the above solution to fix the preparation. Five drops of 
distilled water are then added and the dilute stain allowed to act for 5 
minutes. The smear is then washed with water, dried with filter paper, 
counterstained for 2 or 3 minutes with 0.01% basic fuchsin (Grubler), 
washed, dried in the air and mounted in balsam. 

In specimens thus stained nuclei and cytoplasm are colored red while 
the cytoplasmic granules in neutrophils and endothelial leucocytes are blue. 
The granules of basophils take a distinctive red color. The central portion 
of the eosinophile granules is unstained so that these granules have a very 
characteristic ring-like appearance. The platelets take the red stain faintly. 
The erythrocytes are pink. 

Since the red cytoplasm of lymphocytes is entirely free from bluish 
granules the only differentiation requiring discussion is that between the 
neutrophils and endothelial leucocytes. The granules of the endothelial 
leucocytes are discrete and the cytoplasm is distinctly seen between them, 
while the neutrophilic granules are so thickly placed that little of the cyto- 
plasm can be seen. The neutrophilic granules are larger and more regular 

19 Jour, of A. M. A., Jan. 11, 1908, vol. 1, p. 122. 

20 Arch, of Int. Med. Aug., 1918, xxii, p. 157. 



470 CLINICAL DIAGNOSIS 

in shape than those of the endothelial leucocytes. In pathologic blood in 
which mononuclear myeloblastic cells (neutrophilic myelocytes) are present 
the character of the granules assumes a greater significance. However, 
since the granules of neutrophilic myelocytes and of the so-called meta- 
myelocytes are even more prominent than are those in the polymorphonu- 
clear neutrophils, there is no chance of confusing myelocytes and endothe- 
lial leucocytes, although both are mononuclear. The differential character 
of the endothelial leucocyte is the presence of blue granules in a mononu- 
clear cell. Although the nucleus of this leucocyte frequently has a broken 
outline it does not consist of pyknotic nuclear masses connected by fila- 
ments. The reason that endothelial leukocytes cannot be identified in 
films stained with a polychrome blood stain is that some of these cells are 
entirely devoid of granulation and cannot, therefore, be distinguished from 
lymphocytes when they approach these cells in size. 

A Polychrome Stain for Protozoa. — Mcjunkin's modification of Giemsa's 
stain. 21 This stain is a single solution of polychrome methylene blue, 
methylene blue and eosin. 

The polychrome methylene blue solution is made by measuring accur- 
ately from a buret 50 c.c. of a decinormal solution of sodium carbonate 
into a 500 c.c. beaker. (This is standardized by titration against a standard 
acid using a 1% alcoholic solution of methyl red as indicator.) To the 
carbonate solution is added 1 gm. of methylene blue (Grubler's B. X.) and 
50 c.c. of glycerin (Merck U. S. P.) measured in a 100 c.c. graduate. The 
beaker is placed in a water bath and its contents stirred by a mechanical 
stirrer which is run at the rate of about 200 revolutions a minute. The 
solution is kept at a temperature of from 87 to 89 C. for 1 hour. The 
temperature of the water in the bath outside the beaker should be from 
94° to 96 C. At the end of the hour the beaker is removed from the water 
bath and its contents while still warm poured into a 100 c.c. graduate. The 
beaker is washed out with 5 c.c. of distilled water to recover any carbonate 
which may have precipitated out and this is added to the contents of 
the graduate. 

Into a second graduate is measured enough methyl alcohol (Merck's 
reagent or Kahlbaum's acetone free) to make the total volume of stain 
100 c.c. This methyl alcohol is now poured from the second graduate into 
a 4-ounce bottle and 0.75 gm. of methylene blue (Grubler's B. X.) and 
0.25 gm. of eosin (Grubler's yellowish water soluble) added. The bottle 
is shaken to secure solution of the methylene blue and the eosin in the 
alcohol. After the dyes are completely dissolved in the methyl alcohol 
the polychrome methylene blue solution is poured from the graduate into 
the bottle. The volume of dye is now just 100 c.c. The weights of the 
dyes must be accurate and the glassware free from acid. 

Since some samples of eosin (Grubler's yellowish, water soluble) stain 

21 Jour, of A. M. A., Dec. 18, 1915, vol. lxv, p. 2164. 



THE BLOOD 



471 



the red blood corpuscles a blue that cannot be washed out with water, one 
should use an eosin that has previously been found satisfactory in making 
polychrome staining solutions. 

To demonstrate protozoa and bacteria the smear preparations, which 
may be made on either slides or cover-glasses, may best be fixed in equal 
parts of absolute alcohol and ether for from 10 minutes to a number of hours . 






Fig. 131. — Tube used for diluting serum. 

Distilled water is placed in a staining dish and to this is added 1 drop 
of the stain per cubic centimeter of the water. Cover-glass preparations 
are floated on the solution as soon as the dilution is made and stained for 
from 30 to 60 minutes. Trypanosomes are well stained at the end of half 
an hour while about 1 hour is required to stain Spirochasta pallida heavily. 
The preparation is removed from the stain and an excess of blue is washed 
out with distilled water until the red blood corpuscles are pink in color. 

After washing, the preparation 
is dried in the air and mounted 
in acid-free balsam. Slide prepa- 
rations are best stained in an ob- 
long Petri dish without cover (a 
dish the length and width of a 
slide and about 2 cm. high) , in- 
verted, with one end resting on 
the end of the dish. In this posi- 
tion the specimen will be as near 
the surface of the stain as 
possible. After staining, the slides 
are washed and dried in the air. 
They may be examined directly 
with oil, or they may be mounted 
in acid-free balsam. 

Pappenheims solution of pyronin and methyl green is composed of satur- 
ated aqueous solution of methyl green, 3 to 4 parts, and saturated aqueous 
solution of pyronin, 1 to 1% parts. This stain, which may be used as a 
routine bacterial stain, is useful also in blood work and for distinguishing 
the nucleus of the erythroblasts from its basophilic granules. The nucleus 
and all nuclear fragments stain a beautiful blue and the basophilic granules 
a brilliant red (Morris). The blood spreads should be fixed by heat 
(Ehrlich's Method). 

Ehrlich's Triple Stain. — The words Ehrlich's " triacid " and Ehrlich's 
" triple " stain are often wrongly used as synonyms. The triacid stain, 
a mixture of equal parts of the saturated solutions of indulin, nigrosin, and 




Fig. 132. — Widal test. Field of motile organisms. 
X 900. 



472 CLINICAL DIAGNOSIS 

aurantia, was intended to bring out especially the eosinophile granules. 

It is hard to make up and is now very little used. 

Ehrlich's triple-stain is a mixture of the saturated aqueous solutions 

of methyl green oo, acid fuchsin, and orange G. (Griibler's stains are 

usually used.) The best formula for this stain is that published by Morris. 22 

c.c. 

Saturated aqueous solution of orange G 13.0 

Saturated aqueous solution of acid fuchsin 7.0 

Distilled water 15-° 

Absolute alcohol 15-° 

Saturated aqueous solution methyl green ,. 17.5 

Absolute alcohol 10.0 

Glycerin 10. o 

These fluids are measured in the same graduated cylinder, which should 
not be rinsed out during this procedure and are poured in the order given 

in the formula into a receiving 
flask which should be shaken 
vigorously after the addition of 
each. The methyl green, the 
second portion of alcohol and the 
glycerin should be added drop 
by drop and the flask frequently 
shaken. The mixture can be used 
at once. It seems to improve for 
a time on standing, but later 
spoils. It should never be filtered 
and the bottle containing it 
should never be shaken. The 
blood smear is covered for from 
3 to 20 minutes with a few drops 
Fig. 133.— widai test. Field of agglutinated organ- of the stain obtained on a glass 

rod from as near the center of 
the bottle as possible. It is very difficult to overstain and films suggest- 
ing this usually are underheated. The smear is next washed in 
distilled water, quickly blotted and mounted in balsam. If washed 
quickly in absolute alcohol the granules will stand out more clearly, 
but the nuclei will be paler. In a successfully stained specimen the red 
blood-cells will have a buff color without the slightest shade of red, the 
nuclei of the leucocytes will be a dark green and those of the normoblasts 
almost black, the fine granules will take a lilac stain and the coarse a crim- 
son. This is the only stain which gives a specific color to the fine granules. 
It is for this reason that it was introduced. It is inferior to other stains 
for protoplasm and nuclei and does not in the least stain the Mastzell 
granulation. If one desires to get a good idea of the blood as a whole, other 
1 jour, of A. M. A., Aug. 6, 1910, vol. lv, p. 501. 



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THE BLOOD 473 

stains a so should be used, preferably hematoxylin and eosin, or methylene 
blue and eosin, etc. 

The blood of some persons takes the Ehrlich stain poorly, while that 
of others takes it well. In certain diseases, particularly lymphatic leu- 
kemia, it is almost impossible to get a good specimen with Ehrlich 's stain 
because the basic element is so markedly lacking. 

Ehrlich 's dahlia stain consists of distilled water ioo c.c, saturated 
alcoholic (absolute) dahlia solution 50 c.c. and then, on clearing, 10 to 
12.5 c.c. of glacial acetic acid. The specimen, heated or fixed by alcohol, 
etc., is stained in this solution for from 5 to 10 minutes. 

ERYTHROCYTES 

The red-blood corpuscles are specialized non-nucleated cells which 
consist of hemoglobin, 95%, and of _ stroma. Their chief function is to 
carry oxygen to the tissues and, to a lesser degree, carbon oxide to the lungs. 
In shape they are circular, discoid cells, which in the circulation may 
be cap-shaped (Weidenenreich and Lewis) but which in well-made fresh 
preparation lie flat. In many, but not in all normal cells, a biconcavity is 
apparent but in the secondary anemias especially this is very evident. 
These cells of the normal blood, unless subjected to considerable mechanical 
injury, are perfectly round and in size vary from 6 to 9/x in diameter. 
Those which are not round are called " poikilocytes " (Plate I, 25-28) 
Such cells occur in pernicious anemia especially, e\^en in cases of a mild 
grade and in other anemias of very severe grades especially those due to 
cancer, tuberculosis, etc. They are said to be the results of alterations in 
the plasma, but may also be " badly made " cells. 

Structure. — Red-blood cells are so sensitive that they are about the 
hardest of all cells to study. When fresh they certainly look structureless 
but when stained each method used has indicated a different structure. 
The concensus of opinion now is that all of the fibers, layers, etc., described 
in these cells are artefacts; that the various granule-like bodies seen in 
the fresh cells are not an essential part of the cell and that those seen in 
stained cells are in part, at least, precipitates of the fixing agent or of the 
stain; and that any definite structure, although it certainly must exist, 
is yet to be demonstrated. Ehrlich 's argument that heat must be the best 
fixative agent because it gives homogeneous cells may be the best argument 
against heat. 

Not only their fine but also their coarse structure is in dispute. A true 
cell membrane has never been proven although none doubt that the peri- 
pheral layer of the cell does serve the function of a cell membrane, whether 
it is a membrane or merely a concentration of the stroma at the surface. 23 
or a layer of hemoglobin-stroma in a slightly different physical condition. 
Some still insist that these cells have a true membrane (see page 662). 
23 See Peskind, Am. Jour. Med. Sci., 1904, vol. cxxiv. 



474 CLINICAL DIAGNOSIS 

Those who believe that the nuclei of these cells disappear within the cells 
think it necessary to find some remains of it there. The " nucleoid," so 
often mentioned, is still in dispute; some considering it to be related to 
the nucleus and others to be totally independent. 24 This, also called the 
" differentiated inner body of Lowit," is a nucleus-like structure which in 
stained specimens is very apparent in the center of many of those red- 
blood cells which take a basic stain. It has a fibrillar structure and a cen- 
tral clear space. It may contain an inner body which " may be extruded 
as a platelet . " " This nucleoid develops after- the extrusion of the nucleus ' ' 
said Maximow but Lowit considered it the remains of the now invisible 
nucleus. Whether the erythrocytes in the normal circulations are to be 

considered as living or dead cells is a question 
which recently has attracted considerable at- 
tention. This is largely a question of definition. 
They are very sensitive to injury and degene- 
rate very rapidly when removed from the blood. 
Only perfect cells are seen in the circulation. 
They enter it as such and they leave it before 
the signs of age are apparent (see page 507) . 

Size. — The red cells of the adult vary from 
6 to q/jl (average 7. 5m) in diameter. Hayem 
found that 75% varied from 6.6/x to 8/z; 12.5% 
from 6/x to 6.6/z, and 12.5% from 8 to 911 in 
Fig. 134.— a, b, c, d, four leucocytes diameter. In the normal adult these cells are 

containing Lowit 's organisms (copied r • 1 -r • 1,1 1 c 

from Lowit); e, large granular (and I airly Unit OITTL in Size, although a Very few 
vacuolated?) cell of bone-marrow. -, r 11 r -1 1 11 x , 1 

dwari cells are found at all ages. In the nor- 
mal infant's blood, however, these cells vary much more in size, their 
limits being from 3.3/x to 10.3/x. In disease the adult type of blood 
may assume this infantile condition. There is evidence that the 
red-blood cells of various nationalities differ somewhat, their size 
diminishing as one approaches the equator. In the fresh blood certain 
physiological rhythmical changes in size may be noted, the cells becoming 
somewhat larger in the venous than in the arterial blood (Hamburger). 
Pathologically, they vary much in size. 

Microcytes. — By microcytes is meant a cell under 6/jl in diameter. The 
most measure about 3.5/z and yet some are 2.2/x in size. Not all of these 
small cells are schistocytes, i.e., fragments of larger cells (although one can 
watch the process of constriction of small fragments from red blood-cells 
in fresh blood subjected to mechanical or chemical injury) for microcytes 
are seen in perfect fresh specimens and we find nucleated microblasts 3.5^ 
in diameter which must represent young forms of microcytes . Microcytes 
are not biconcave as a rule but are spherical and hence have a deep color. 
They occur normally in large numbers in the embryo infant and especially 

24 Maximow, Arch. f. Anat. u. Physoil, 1899. 





THE BLOOD 475 

in the blood of premature children in which cases they are often polychro- 
matophilic. They are found rarely in the healthy adult, but are common 
in all anemias, especially the primary and severe secondary anemias. 

Macrocyte is a term applied to cells from 9 to 1 2/z and above in diameter; 
for cells from 12 to 16/z the term megalocyte is used, and for those above 16 /j, 
gigantocyte. These cells occur in largest numbers in pernicious anemia. 
Some believe that if 10% of the red cells of any given case are macrocytes 
a diagnosis of pernicious anemia is justified. They also occur in leukemia 
and in chlorosis. In chlorosis they are often very pale, hence are termed 
" chlorotic "or " dropsical " cells. They are 
common in cases with cholemia, which is of 
interest since patients with pernicious anemia 
are so often jaundiced (Osier). Their large size 
may be due to hydremia for it is well known 
that in hydremia the plasma is quite con- 
stant in its water-content and that the 
variations in the water of the total blood 
affect especially the cells. In pernicious ane- 
mia the largest cells are some times the darkest f lG - 135.— The intestine of an m- 

fected mosquito with oocysts at- 

and some of the microcytes are exceedingly tached. (From Braun.) 

pale , while in secondary anemia the reverse is true (see plate I). 

Staining Properties. — Red blood-cells like all other cells while "alive" 
are achromatophilic . If fixed by an agent which prevents post-mortem 
changes all that are normal are monochromatophilic and, since they 
take from a mixture only acid stains, are acidophilic . Since it is the he- 
moglobin that takes the stain, the amount of this pigment may be esti- 
mated from the depth of their color. 

Red cells which either as a whole or in part take other than the acid 
component of a stain are termed polychromatophilic or basophilic. Under 
this term we do not now include the basophilic granules to be described 
later. Eosin stains basophilic red cells more faintly than normal and if 
followed by a basic stain, such as hematoxylin, will be supplanted by it. 
Basophilic corpuscles are usually larger than the normal have less bicon- 
cavity and often are poikilocytes. Stained with hematoxylin and eosin 
such cells take a violet tint ; with Ehrlich's stain a fainter orange than nor- 
mal, or a grayish color; with polychrome methylene blue they stain a 
bluish violet color. 

Ehrlich explains basophilia as a coagulative necrosis, an " anemic 
degeneration." In favor of this view are, that other signs of degeneration 
also are present ; that it can be produced in animals by inanition ; that these 
cells appear within 24 hours after a hemorrhage, that is, before nucleated 
cells or other signs of regeneration have appeared; and that basophilia 
affect especially the megaloblasts and other red cells which are abnormal 
in size or shape. Another view is that the basophilic cells are young since 



476 



CLINICAL DIAGNOSIS 



young cells certainly are basophilic. They are met with in pernicious 
anemia, in the grave secondary anemias, especially those due to cancer, 
in the eruptive fevers, malaria, the purpuras and after various blood poisons. 
Other cells are " fuchsinophilic " (Plate I, 35); that is, if stained with 
Ehrlich's triple stain they are too red. Since so many of the nucleated reds 
of the bone marrow are fuchsinophilic this also is considered a sign of a 
young cell. These cells also are usually distorted, as if very soft. The same 
is true of the basophilic cells and nearly all nucleated red cells are slightly 
basophilic. Basophilia and fuchsinophilia however are not the same. With 
the blood stains now in common use we can disregard fuchsinophilia but 

are even more confused by baso- 
philia since methylene blue, the 
basic stain used, is exceedingly un- 
trustworthy. 

Using the best of stains we may 
say that young red cells in general are 
basophilic and that many degene- 
rating cells become so. For this 
reason it is hard to say whether micro- 
cytes, macrocytes and the basophilic 
granules are signs of regeneration or 
degeneration, but it is improbable 
that they have always the same sig- 
nificance. Theobald Smith, in des- 
cribing a case of purpura, first 
£wo 3 ^^ suggested that basophilia is evidence 

il^^^^^^t^ys^.^^oi^yaaih of cells and later em- 
vivax). (From Braun.) phasized this in his studies on Texas 

fever. 25 Walker (see p. 862) found basophilic cells in the normal blood of all 
lower vertebrates, while in the blood of the fetus of the dog and guinea-pig 
there were even 90 times as many such as in the blood of the mother. In nor- 
mal human marrow the basophilia of a cell is in inverse proportion to the 
amount of hemoglobin which it seems to contain hence the term "anemic 
degeneration"; but this lack of hemoglobin could be primary as well as 
secondary. 26 Walker suggests that these are "cells hurried into the circu- 
lation while too young " and therefore are as good and certainly a much 
more convenient index of anemia than is the blood-count. 

Partial polychromatophilia is best illustrated by the definitely basophilic 
areas of Maragliano's endoglobular "degeneration" (see p. 448). The 
probability is that many of the so-called " inner bodies," " nucleoids," and 
other so-called evidence of cell-structure are nothing but these areas of 
changed protoplasm. Sometimes the areas markedly resemble malarial 

25 Walker, loc. cit. 

26 See also Stengel, Contrib. from Pepper Lab., Univ. of Pa., 1900. 




THE BLOOD 



477 




Fig. 



137. — Attitude of mosquitoes on wall, a, Anopheles; 
b, Culex. 



parasites, while others when extruded resemble platelets. These degenera- 
tions certainly led to many mistakes in the diagnoses of malaria before the 
chromatin-staining mixtures were used. 

The ring bodies described by Cabot 27 in some of the red cells of anemic 
blood and which require for their demonstration the polychrome methylene 
blue-eosin mixtures, he suggests are nuclear remains. These appear as 
rings, ovals, or bands and ap- 
parently are not related to the 
basophilic stippling. They occur 
especially in pernicious anemia 
but also in the leukemias and in 
various secondary anemias. 

In specimens heated too 
quickly many of the cells have at 
their periphery a row of large 
dots which are not true granules. 
In certain cases of malaria 
(those we have seen all have 
been tertian and from the 
Tropics) the infected cells show 
a remarkable granulation (Plate III, 10, 13). These granules are of 
quite uniform size about i/x in diameter. They can be seen in the fresh 
unstained cell (the lead granules cannot). They stain purple in the 
Hastings' stain, while the rest of the cell stains paler than normal, in 
fact may be almost colorless as if the hemoglobin had been condensed 

into these dots. We have seen 
red cells in which these granules 
appeared suspended in a hyaline 
envelope around the parasite. 

The " methylene blue degene- 
ration of Ehrlich " is the name 
given to a beautiful blue mesh- 
like fibrillation of red cells in 
specimens of fresh blood stained 
by this dye. 

Vital Blood Staining. — To 
a * study the granules and fibers of 

FIG. i 3 8.-Heads of mosqmtoes. a, Culex; 6, Anopheles. unfixed ^ ^ dropg Qn ^ 

smear before it dries a granule of methylene blue or neutral red and then 
immediately seals the cover glass to the slide with paraffin. Beautiful 
threads of fine granules are soon seen. Another excellent method of vital 
blood staining was that used by Rosin 28 who covered a cover-glass 





27 Jour, of Med. Research, 1903, vol. lv., p. 15. 

28 'Rosin and Bibergeil, Zeitschr. f. klin. Med., 1904, vol. liv, p. 



107. 



478 CLINICAL DIAGNOSIS 

lightly with a saturated alcoholic solution of methylazur or of toluidirj 
blue, which he then allowed to dry, and made the blood smear on this 
stained surface which was then at once inverted over a hollow slide 
with vaselined rim. The changes in blood thus prepared can be studied 
for even 24 hours. 

Vogel and McCurdy 29 recommended the following method : 

One saturates 0.85% salt solution with 
brilliant cresyl blue (see below.) This is fil- 
tered through^a double paper to take out 
the excess of dye substance and to prevent 
precipitation on the slide. It is better to 
centrifugalize the stain before using it in 
order to be sure that no undissolved parti- 
cles remain in suspension. If any precipi- 
I ill tate is present in the stain it later will be 

Fig. i40.-Pilaria bancrofti. X 50- thiwn down when cen trifugalizing the 

mixture of blood and stain and will cause confusion when counting the 
reticulated cells. 

The stain should be freshly prepared for use as follows : Saturated solu- 
tion of brilliant cresyl blue in 0.85% salt solution and 

Salt solution, 0.85% aa 5 c.c. 

Sodium oxalate solution, 2% 2 c.c. 

Add the oxalate to the salt solution, then mix with the stain and filter. 



Fig. 141. — Microfilaria bancrofti with sheath, from blood taken at 

Charleston, S. C. Stained with hematoxylin. Travis, U. S. Public 

Health Service 

The finger is so punctured that we can get a free flow of blood. A good 
sized drop is drawn into a red cell-counting pipette using the stain as a 
diluent. After thorough mixing, this is allowed to stand for 10 minutes 
and the contents of the mixing chamber then blown into a centrifuge tube 
and centrifugalized. * The staining fluid is drawn off with a capillary pipette 

29 Arch, of Int. Med., Dec, 1913, xii, p. 707. 



THE BLOOD 



479 




until only the sediment of cells remains, and these are then drawn from the 
bottom of the centrifuge tube in a capillary pipette. A drop of this fluid 
is placed on the end of a clean slide which has been slightly warmed in a 
flame and spread as in making ordinary blood-smears. Beautiful threads 
of fine granules are soon seen in certain cells. The preparation will keep 
indefinitely if mounted in neutral balsam or damar and not exposed to 
strong daylight. 

In counting the cells an Ehrlich eyepiece is of 
great assistance. In suitably stained specimens 
the granulo-filamentous or reticulo-filamentous sub- 
stance appears in the form of granular particles 
which are sometimes discrete but more often form 
threads which frequently are woven into skeins or 
wreaths of great complexity and which fill a con- 
siderable portion of the cells. In the blood of in- 
fants these reticulations are found in from 5 to 
10% of the erythrocytes and in normal adult blood 
in from 0.5 to 2%. In severe anemias, however, 
their number may run as high as 18 or 20% and in 
hemolytic jaundice, where they are most numerous, 
they may occur in still greater proportions. 

Vogel and McCurdy conclude that this granulo- 
filamentous substance is not derived from the 
nucleus, is different from polychromatophilia and 
from the basophilic stippling seen in fixed prepara- 
tions stained with panchromatic dyes, is not a 
preformed structure but a precipitation product of 
the stain and is an evidence of youth of the cells and 
not of degeneration. In conditions in which a severe 
drain on the erythrocytes is being sustained by a 
well-functionating bone-marrow, large numbers of 
reticulated cells are found, whereas in aplastic cases 
they may be diminished almost to the point of ab- 
sence. That is, the reticulated cells, in a manner 
somewhat comparable to the behavior of the ery- 
throblasts, afford a direct insight into the hemato- 
poietic activities of the bone-marrow. For clinical 
purposes they form a more convenient measure of 
this function than do the nucleated cells as their percentage relations to 
the erythrocytes can be more easily and accurately determined and their 
enumeration is to be urged as a part of the study of the blood in all cases 
of severe anemia, 

The Basophilic Granulation of Grawitz (Plate II, 22, 24, 25). — In 
certain conditions, especially lead poisoning, pernicious anemia, leukemia, 



Fig. 142. — Mature larva 
escaping from proboscis of 
Culex fatigans. One mature 
larva coiled in base of pro- 
boscis. From mosquito dis- 
sected after being infected 
at Charleston, S. C. U. S. 
Public Health Service. 






480 CLINICAL DIAGNOSIS 

etc, certain of the red-blood cells when stained with any basic stain, but 
particularly with gentian violet or methylene blue, are seen to contain 
minute granules. These granules are not visible in fresh unstained speci- 
mens and do not increase in specimens on standing. 

They are best demonstrated as follows : The air-dried smears are fixed 
for from 3 to 5 minutes in absolute alcohol, washed in water and, while 










FlG. 143. — The spirochete of relapsing fever. X 1200. 

still wet, are stained for a few seconds or much longer with Lofner's methy- 
lene blue. They are dried, or examined, while still wet. The bluish-black 
granules will stand out against the clear green corpuscles. 

Pappenheim proposed a stain intended to differentiate them from nuclear fragments. 
Stain I. Acid, carbol. liquefact., 0.25; aqua dest. 100; methylene green (pur.), 1. 
Stain II. Acid, carbol. liquefact., 0.25; aqua dest., 100; pyronin (pur. ), 1. 

Fifteen cubic centimeters of I and 35 c.c. of II are well mixed and filtered. The 
blood-smear fixed by heat (not alcohol) is stained for a few seconds with this filtrate. 
Nuclear fragments will stain a deep greenish-blue color, Grawitz's granules a bright red. 

When numerous, 5 or 6 of these "stippled cells " may be seen in a field, 
but as a rule several fields must be searched in order to find 1 such cell. 
The cell may contain but 1 or a few of these granules, but as a rule it is well 
sprinkled even to such a degree that some are almost uniformly blue. The 
granules vary from dust-like size to ijuor more in diameter. They may 
occupy any part of the cell, but are uniformly distributed as a rule and 
(many think) are situated in the external layers of the protoplasm. Stip- 



THE BLOOD 



481 






pled cells are met with in the severest anemias, especially the primary 
pernicious (in which they are large and conspicuous) and in secondary 
anemia especially that due to cancer of the gastrointestinal tract; in 
cachexia; in leukemia, in which cases they are not numerous; in septic 
processes; and in chlorosis (although some say they are rare, others as 
Stengel and Pepper who found them in 2 of 1 8 cases,say they are common) ; 
in phthisis, especially after the secondary infections develop; in lues; chronic 
parenchymatous nephritis; small contracted kidney; cirrhosis of the liver 
(Grawitz) ; in gout (in which condition they may be numerous, especially 
in cases with hematoporphyrinuria, and yet in other forms of arthritis 
with even severe blood changes they are very rare); typhoid fever; pneu- 
monia ; lues ; nephritis ; etc. Guyot found them regularly in a case of hemo- 
globinuria due to cold. 
They are found in the 
blood of Europeans who 
recently have moved to 
the Tropics. 

But that condition 
in which they are parti- 
cularly important and 
numerous is lead pois- 
oning, since no other 
blood changes need be 
present. Some claim 
that they are present 
in the blood of all lead- workers, but they certainly cannot always be 
found in the time at the disposal of the ordinary clinical worker. They 
vary much in number from day to day. As a rule they appear early, even 
after but 4 days' exposure to lead, and they may persist in the blood for 
over 20 years. Since these granules may be the first sign of an anemia we 
may possibly diagnose this condition even before the count drops. They 
also are the last sign to disappear. 

Grawitz interpreted these granules as areas of coagulated necrosis, 
hence the name " Grawitz basophilic granular degeneration," and Ham- 
el, 30 White and Pepper, 31 Stengel and Pepper, 32 Bloch and others agree. 

On the other hand similar granules are found in the blood of the embryo, 
and nucleated reds often contain them, which is good evidence that some 
at least are not nuclear fragments and that they bear some relation to blood 
regeneration. 

Cadwalader 33 distinguished 3 groups of granulated red cells : those 

with the granules in fine and coarse thread-like strands, found sometimes 

30 Deutsch. Arch. f. klin. med., May 23, 1900. 

31 Am. Jour. Med. Sci., Sept., 1901. 

32 Am. Jour. Med. Sci., May, 1902. 

33 Bull, of the Ayer Clin. Lab., Univ. of Pa., January, 1905. 

31 



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Fig. 144. — Colloidal gold curves. 



482 CLINICAL DIAGNOSIS 

in normal blood; those with fine dot-like granulations, the most common 
form, especially in lead poisoning and pernicious anemia; and those with 
dense coarse basophilic masses seen in cases of lead poisoning in which 
nucleated reds are plentiful and suggesting by their position and size frag- 
ments of a nucleus, although the transitional stages between fragmenting 
nuclei and these granules are not found and they are rare in the bone- 
marrow where karyorrhexis is most common. 





■ j|r- 




■ ; - • ; * 

& . ■ ' 

..w. _ » ,_, 



Fig. 145. — 'Smear of the spinal fluid Fig. 146. — -Smear of the spinal 

of a case of epidemic cerebrospinal fluid of a case of meningitis due to 

meningitis. Diplococcus lanceolatus. 

At this point we take the liberty to state that practically every basophile granule 
found in red blood-cells has been described by one or another writer under this one title. 
In our opinion there are at least 3 different basophilic granulations in red blood-cells, 
which have, we suspect, no relationship the one to the others. 

The granules seen in lead poisoning are not visible in fresh blood; they are seen 
also in normoblasts with perfect nuclei. These are always accompanied by other evi- 
dences of blood regeneration. The granules seen in malaria are described on page 668. 
They can be seen in the fresh cell. Compared side by side with the Grawitz granules 
they do not look at all alike. The granules described by Vaughan (see page 449) as 
remnants of nuclei do not resemble the Grawitz granules. We judge from Cadwalader's 
figures that he includes them as his coarse variety. 

Naegeli 34 considered them related to blood regeneration and Boellke 35, 
denied that they bear any relation to the nucleus. 

Number of Red Blood-cells. — The average count of the red blood-cells 
of the normal adult man is usually given as 5,000,000 per cubic millimeter 
of blood; of the woman, 4,500,000. In a healthy young man, however, 
it is more common for the count to vary from 5,000,000 to 6,000,000. 

By polycythemia is meant a condition with more cells than this per 
cubic millimeter ; by oligocythemia, one with a smaller number. It is evident 
that such numbers are simply relative, that variations in the count may be 

34 Munch, med. Wochenschr., 1904, No. 5. 
85 Virch. Arch., 1904, vol. clxxvi, S. 47. 



THE BLOOD 483 

due either to actual variations in the total number of red blood-cells in 
the body or to variations in the amount of plasma. 

The blood-count may vary in different parts of the body. Oliver 36 
found that anything which increases the blood-pressure locally causes a 
rise in the count at that point. For instance, the count is higher in a limb 
that has been hanging in a dependent position and in areas subjected to 
active or passive motion. 

Exercise of a part will raise the count locally and the application of 
cold and of heat will lower it or raise it, according as it produces stasis, 
vasodilatation, or constriction. 

Excessive exercise, Willebrand found, would raise the general count 
of red cells from 3 to 23% (average 12.3%), and that of leucocytes from 
19 to 97% (average 47%)- 

Physiological Variations. — The sex variations have already been 
mentioned. This difference is present only during the menstrual period 
of life, since the count of girls until their fifteenth year averages 5,444,000 
and for boys of the same age 5,102,000; also between the ages of 40 and 60 
the count of women averages 5,000,000. 

The red cell count varies much with age. The maximum is at birth, 
in which case it may be even 7,000,000 but as a rule is lower — e.g., 5,740,000 
(Stengel and White). Otto found the average for the first 4 days to be 
6,155,000; in 1 child 10 hours old it was 6,910,000. After the first 4 days 
of birth the count begins to drop and reaches a minimum in about 1 year 
(see below). This count depends somewhat on the time at which the 
umbilical cord is tied, since tying it off late may result in a gain of almost 
1,000,000 cells per cubic millimeter. These high counts at birth are due 
probably to an unusual concentration of the blood resulting from the loss 
of water. They last but for a few days, not over 10, after which time the 
nucleated reds also disappear. 

From birth until about the tenth year the count is relatively low and 
then slowly rises. There is considerable difference of opinion as to the 
exact age when it reaches its minimum. It rises from the time of puberty 
to 30 years of age, during which period the count in young healthy persons 
often ranges from 5,500,000 to 6,000,000 cells. Between 30 and 50 years 
a count of 5,000,000 for men and 4,500,000 for women may be considered 
normal. After 40 the count is inclined to drop slowly in men and to rise 
in women. 

Not satisfied with the age curves usually quoted in text-books, we have tabulated 
the counts of all patients with apparently normal blood and those of our medical students 
(see page 444). 

We have calculated means, not averages, since the extremes are always subject to 
criticism and should be reported independently. For a discussion of these low hemo- 
globin estimations see page 444. 

36 Brit. Med. Jour., 1896. 



484 



CLINICAL DIAGNOSIS 



Blood of fatients 

MALES 



Years 


Cases 


Reds (mean) 


Hb mean 
Per cent. 


Index 


Leucocytes 


6 to 15 


5 


5,560,000 


85 




7500 


16 to 25 


36 


5,200,000 


85 


0.8 


6500 


26 to 35 


69 


5,300,000 


90 


0.85 


7000 


36 to 45 


42 


5,500,000 


90 


0.82 


5500 


46 to 55 


21 


5,300,000 


80 


0.75 


9000 


56 to 65 


9 


5,000,000 


80 


0.8' 




66 and over 


5 


4,000,000 


60 


0.77 


7500 



10 to 15 


5 


5,000,000 


75 


0-75 


8000 


16 to 25 


43 


4,500,000 


77 


0.85 


7500 


26 to 35 


55 


4,500,000 


80 


0.88 


7200 


36 to 45 


34 


4,600,000 


72 


0.80 


7700 


46 to 55 


17 


4,500,000 


77 


0.85 


7000 


56 to 65 


10 


4,500,000 


70 


0.78 


6000 


66 and over 


3 


4,700,000 


65 


0.7 


7000 



A study of the blood of 176 men students from 20 to 25 years of age gave the follow- 
ing: Means of reds, 5,000,000 (extremes 4,500,000 and 6,700,000); 14 (8%) were below 
5,000,000 and 15 (8.5%) above 6,000,000. Mean of leucocytes, 7500 (52 cases); of hemo- 
globin, 14,5 gms. (Miescher), 92% (Fleischl), 95% (Dare), 92% (Gowers). 

Women medical students, same age limits, 16 cases. Mean of red 4,800,000; of 
leucocytes, 8000; of hemoglobin, 11 gms. (Miescher), 85% (Fleischl), 87% (Dare), 
82% (Gowers). 

Nutritional Conditions. — The red cell count of thin muscular persons is 
somewhat higher than that of fleshy persons. A large meal may be followed 
by a temporary slight decrease, said to be due to an increase in the plasma. 
During hunger periods the count often rises a half million cells in 24 hours. 
This is attributed to concentration of the blood. 

The temperature has an influence on the count. In winter the counts 
averaged about 500,000 cells per cubic millimeter more than in summer 
(this was well seen in some of our students' counts). The change of resi- 
dence from temperate zones to the Tropics may lead to a drop in the count 
of from 500,000 to 2,000,000 cells. 

Pregnancy. — The counts of pregnant women and also of the fetuses are 
said to drop during the last part of pregnancy; that of the mother about 
half a million cells and the hemoglobin 20%. The count of the fetuses of from 
7 l A to 8% months was found to be 7,000,000 and at 9 months 6,500,000 
(Biondi and Gardini). The bloods of mother and child are independent 
enough so that if the mother has an anemia-producing disease the child 
can preserve its count fairly well, and vice-versa. 

Thompson made a very careful study of the bloods of 12 pregnant 
women in Dr. J. Whitridge William's clinic. He found that the counts 
and the hemoglobin decreased slightly from the fourth to the eighth month 
and then rose to normal at term. The curve of the specific gravity of the 
blood ran in general parallel to that of the red cells and hemoglobin, but 
the initial fall and the terminal rise were more accentuated than of those 
curves. The minimum figures (1.0408) were during the sixth month. 



THE BLOOD 485 

Altitude. — That the red cell count rises as persons ascend to high alti- 
tudes is a phenomenon long ago witnessed but not yet thoroughly explained. 
The rise amounts to about 50,000 cells per 1,000 feet. The count returns 
to normal as they descend, or at the latest 36 hours after. The rise in the 
count is especially marked if the ascent to a considerable height is sudden. 
It is slight following an ascent of 1200 meters, slight and tardy after an 
ascent of 1800 meters, but it is immediate and considerable after an ascent 
of 3000 meters. This increase in the count is certainly too rapid to be 
explained by new blood formation and after the descent there are no signs 
of blood destruction. The rise is most marked in invalids, especially those 
with lung tuberculosis. The symptoms of anemia are even aggravated 
by the ascent. 

There would seem to be 2 factors which raise the blood count in these 
cases, a temporary change in the 
distribution of the blood-cells and 
later, in 8 or 10 days, an increased 
production of new cells. 

Miescher and his pupils, as a re- 
sult of work with animals, claimed 
to demonstrate that a diminished 
oxygen tension is a stimulus to new 
blood formation. Later, practically 
all of this experimental work was 

r Fig. 147. — -Smear of the spinal fluid of a case of 

Challenged. Certainly animals Sent meningitis due to Bacillus influenzas. 

to high altitudes have given disappointing results. 

Others (Grawitz) say that the blood is concentrated because of evapora- 
tion of body water, and yet the solids of the plasma do not change as much 
as does the count; others (Truntz) that it is due to an accumulation of 
cells in the capillaries; that the cells lived longer; that there is at first a 
fragmentation of the red cells which explains the early increase and then 
a true new formation follows (Koppe) ; finally that there is a peripheral 
vasoconstriction which causes an increase in the tissue lymph and so a 
concentration of the blood (Bunge) . One of the best papers on this subject 
is that of Campbell and Hoagland 37 who consider that the rise in the count 
is explained largely as a change in the distribution of the blood cells which 
results from the lowering of the blood pressure and is due to the lowered 
barometric pressure and to a compensatory increase in the heart action, 
the increase in pulse rate being almost parallel to that of the count. Mosso 
showed that there is at high altitudes a peripheral vasodilatation, hence a 
stasis in the dilated capillaries. If a person remains at the high altitude 
the heart will soon compensate for all these factors and the count return 
to normal. The difference in temperature has some, some say the most, 
influence on the blood count, hence at Colorado Springs it is about 

37 Am. Jour. Med. Sci., November, 1901. 




486 CLINICAL DIAGNOSIS 

800,000 cells per cubic millimeter lower than at the City of Mexico, although 
the 2 places have the same elevation. Experiments with rabbits showed a 
decreased count in the mesenteric circulation corresponding to the rise in 
that of the peripheral circulation. 

The observations of Gaule, who studied his blood during a balloon 
ascension and found that the rise was accompanied by the appearance of 
many nucleated reds, have not been confirmed. 

r 




Pig. 148. — Cells from a pleural exudate. X 900. 

Drugs and Therapeutic Measures. — Among the drugs which increase 
the count of the red cells are iron, which is almost a specific remedy in 
chlorosis, and arsenic, a drug equally valuable in pernicious anemia. 
Mercury in large doses causes an anemia. The destruction of injured cells 
by this drug may explain the Justus test for lues (see page 654). Lead 
causes a chlorotic anemia which some attribute to a direct injury of the 
red blood-cells (see page 481). 

Any drugs which can change the amount of plasma by causing rapid 
losses of fluid to the body, as diuretics, emetics, purgatives, diaphoretics, 
may cause a temporary rise in the count; but this is hard to demonstrate 
in the wards. 



THE BLOOD 487 

Cold baths in typhoid fever cause an average increase of 1,860,000 in 
the red cell-count and an increase of the specific gravity of the blood 
(Thayer) which disappears in about 1 hour. This is due to a temporary 
stasis in the capillaries. Breitenstein thinks the effect of a cold bath is 
greater in a typhoid patient than in a normal man since the distribution 
of cells in the former is abnormal before the bath. 

After operations there is often a transitory rise of from 100,000 to 
1,000,000 cells, probably due to changes in the peripheral circulation. 

Pathological Conditions. — The toxins of certain of the specific fevers 
often cause a marked anemia. This may be due to a definite destruction 
of red cells, in which cases there is an increase of the pigment in the urine, 
and sometimes, although rarely, to definite minute or larger hemorrhages. 
In other cases changes in the bone-marrow may be important. 

Chronic Cachexia. — The toxins of the more chronic infections which 
produce chronic cachexia may produce a definite anemia. Of these, tuber- 
culosis, cancer and lues are the best illustrations (see pages 634, 648, and 
652). The methemoglobin-producing poisons may diminish the count 
because of their direct destruction of the cells. Among these are pyro- 
gallic acid, the chlorates and certain of the coal-tar products, as anti- 
febrin and phenacetin. 

Polycythemia. — In this section we follow closely the excellent article 
by Watson-Wemyss. 38 Polycythemia, an increase in the number of red 
corpuscles per cubic millimeter of blood, may be transient or permanent. 
The transitory polycythemia is usually due to a concentration of the blood 
caused by loss of fluid from the plasma; to local cyanosis, to poisoning 
with acute phosphorus (in which case the count may be even 8,650,000) 
and carbon monoxide (the count even 6,630,000). Vomiting alone cannot 
explain these high counts. 

Permanent or absolute polycythemia would seem to be the result of 
an absolute increase in the production of red cells. In favor of this view 
are the signs of active blood formation (nucleated red cells, polycnromato- 
philia, leucocytosis, eosinophilia, etc.). This polycythemia may be a 
secondary process (erythrocytosis) or a primary process (erythremia) . 

Erythrocytosis, or a permanent polycythemia, the cause of which is in 
part at least understood, may be due to high altitudes; heart diseases, 
especially the congenital forms in which the count may vary from 8,000,000 
to 9, 000, 000 per c.mms. ; also mitral-valve disease and adherent pericardium, 
lung diseases, especially empyema, acute miliary tuberculosis and pneu- 
monia; chronic stimulation of the bone-marrow by poisons, as phosphorus, 
acetanilide, etc. 

Erythremia is considered a primary disorder since we cannot ascribe 
it to any known cause. It seems due to abnormally increased activity 
of the marrow and is often accompanied by enlargement of the spleen, 

38 Edinb. M. J., Feb., 191 1, vol. lv, N. S., p. 129. 



488 CLINICAL DIAGNOSIS 

cyanosis and sometimes by arterial hypertension. This condition has 
been named Vaquez's disease, Osier's 39 disease, splenomegalic polycy- 
themia, etc. The red cell-count of these cases varies from 6,000,000 to 
13,000,000. The red cells are of normal size, nucleated red cells are not 
rare and poikilocytes and polychromatophilic cells are sometimes seen. 
A polymorphonuclear neutrophile leucocytosis (from 20,000 to 91,000) is 
the rule although a leucopenia has been found. There is also an absolute 
eosinophilia. Myelocytes are rarely found. 

Osier reviewed 9 cases, 4 of which he reported. The cyanosis was 
extreme and lasted even for years; the highest count was Koester's of 
13,600,000 and but 1 was below 9,000,000 (8,250,000); the hemoglobin 
ranged from 120 to 150%, the specific gravity from 1.067 to 1.080 and the 
leucocytes from 4000 to 20,000 (the most of the counts were below 10,000). 
Zamfirescu reported this condition in a woman with polycythemia, cyano- 
sis, dyspnea and cough. Kikuchi one with bronchiectasis. Turk 40 reported 
7 cases like Osier's, 2 with autopsy, the counts of which varied from 
7,700,000 to 10,600,000. He suggested that erythremia is due to a primary 
hyperplasia with increased function of the erythroblastic myelogenous 
tissue and so is analogous to leukemia. Other interesting cases of poly- 
cythemia without cyanosis have been reported, as Zandy's 41 who proposes 
the term " erythrocytosis." Turk 42 mentioned cases characterized merely 
by the high count. 

In this connection we should remember that we usually count capillary 
and not arterial or venous blood and that the count in the capillaries may 
not be the same as that in the vessels although it usually is the same as 
that in the veins. To appreciate this important point the student should 
study the capillary field of a frog's mesentery or a rabbit's ear. He will 
see some wide capillaries, some through which the corpuscles pass in single 
file and others so narrow that the cells do not enter them at all. If now an 
active congestion or a venous stasis be produced the capillaries fill up with 
cells, even those which before transmitted only plasma. In this local area 
a count of capillary blood certainly would be higher than before. 

Local changes in counts are much more marked in the case of the 
leucocytes than of the reds since the former collect in the smaller vessels, 
forming layers along the walls. 

Local cyanosis may deceive one much; for example, some cases during 
life have normal blood-counts who at autopsy show a condition suggesting 
pernicious anemia. The same is true of certain dysenteries. 

High counts are met with following the use of various coal-tar products. 

One of our students recently became cyanotic after handling aniline 

oil. His red cells then were 5,900,000, hemoglobin 107% (Dare) and leu- 

39 Am. Jour. Med. Sci., 1903, vol. cxxxvi. 

40 Wien. klin. Wochenschr., 1904, Nos. 6 and 7. 

41 Miinc. med. Wochenschr., 1904, No. 2j. 

42 Deutsch. med. Wochenschr., 1904, No. 20. 



THE BLOOD 



489 



oocytes 6100. Six days later the red count was 5,084,000 and the hemo- 
globin 78% (Dare). 

Resistance of the Red Blood-cells. — Many methods have been proposed 
for determining the resistance of the red blood-cells in the hope of explaining 
such phenomena as hemoglobinemia, etc. At first the methods used were 
mechanical, such as shaking, chemical and electrical, but now they are 
biological. 

Hamburger's Method. — Formerly the resistance of the red cells was 




3 Fig. 149. — 'Fatty acid crystals from the contents of an ovarian cyst. X 400. 

tested by exposing the washed red cells to salt solutions of various strengths 
to determine the concentration at which hemolysis takes place. In 16 
small test-tubes are mixed i drop of a suspension of the washed corpuscles 
and 1 c.c. of a series of sodium chloride solutions, the lowest of which is an 
0.4 % solution and each succeeding one 0.03% higher than the preceding one. 
The tubes, after a gentle shaking, are allowed to stand 6 hours. Normal 
blood plasma is isotonic with an 0.9% NaCl solution, but normal corpuscles 
are not laked unless the concentration is less than about 0.4%. In con- 
genital family cholemia the hemolysis may begin at 0.7 or even at 0.9%. 
Stengel diluted the blood 1 ,in a Zeiss leucocyte pipette, to 10, with sodium 
chloride solutions varying from 0.42 to 0.52% shaking the mixtures well. 
The blood is then blown into small tubes sealed at one end. These after 



490 CLINICAL DIAGNOSIS 

standing are centrifugalized for from 2 to 5 minutes, then held against a 
white paper to determine the presence of hemolysis. Stengel found that 
saturation with an excess of carbon dioxide (as in congestion) causes no 
morphological alteration in the cells and yet increases their vulnerability; 
that cold produces marked changes ; and that hypotonic salt solutions have 
the power in the test-tube as well as within the blood vessels of decolorizing 
and vacuolating the red blood-corpuscles. (This point is disputed by many 
workers, who find that even the intravenous injection of distilled water 
does not lake any blood-cells) . Heat of even slight degree changes the shape 
and size of the corpuscles and finally decolorizes them. A higher degree 
causes budding, vacuolation and, a somewhat higher degree, complete 
fragmentation (see page 446). 

Mechanical Influences. — In some conditions the cells have been 
found to have decreased resistance to shaking. Meltzer has shown 43 that 

the effect of shaking depends upon the 
rapidity of vibration, that for each blood 
there is a minimum and maximum 
rate which the cells can bear without 
destruction. Laker tested the cells by 
passing through their suspension the 
discharges of a Ley den jar. Various 
other methods have been proposed, but 
with as yet little result. For the 

Fig. iso.-Cholesterol crystals. X 400. much mQre important BIOL OGICAL 

methods see page 588. 

Color Index. — By color index is meant the percentage of hemoglobin 
divided by the percentage of the red blood-cells. This figure, as Duncan 
first showed, has considerable value. For the denominator, 5,000,000 red 
blood-corpuscles is considered 100%, while the numerator is the per cent. 
of hemoglobin read with any good instrument. The color index is less than 
1 in practically all secondary anemias. It is especially low in chlorosis, 
averaging about 0.5, and in some cases reaching as low as 0.3 . In pernicious 
anemia, on the other hand, the index is increased, averaging about 1.04 
(Cabot) and in 1 case reaching 1.75 (count 1,000,000, hemoglobin 35). 
The high color index is of value in differentiating pernicious anemia from 
certain cases of latent cancer of the stomach, a diagnosis often hard to 
make. The color index is not a strict mathematical calculation: the 2 
figures are too approximate for that. Five million is only approximately 
the normal red-cell count and few hemoglobinmeters read normal blood 
at 100%. As stated on page 484 we found, taking instruments as they 
come, that the index in normal persons varies from 0.80 to 0.88. The color 
index in the case of our male students, using the Fleischl and Gowers 
instruments, was 0.84 and with Dare, 0.87. The index of the blood of our 

43 Johns Hopkins Hosp. Rep., vol. ix. 





. /---. 


ps^ 


; • xy - 


/ ^^ 


:S 


■ 1 
1 h j 

1 


s^/^**"*-^ 


. . _:_ j 



THE BLOOD 491 

women medical students using the Fleischl was 0.88, the Dare, 0.9 and the 
Gowers, 0.82. Yet we often see persons with the above figures reported 
as cases of " mild chlorotic anemia." 

We therefore approached the question in still another way. Among 
our students' reports were 53 records of counts and Miescher hemoglobin 
estimations made on the same normal bloods at the same hour. The counts 
varied from 4,600,000 to 6,700,000, and the hemoglobin from 10.9 to 17.2 
gms. per 100 c.c. If in each case the number of grams of hemoglobin per 
1,000,000 cells be reckoned, the mean of these figures should be an approx- 
imately normal color index. These quotients fell within surprisingly close 
limits since 42 of the 53 varied from 2.2 to 2.8 gms., while the mean was 
2. 03 gms. per 1,000,000 cells. We believe this standard of hemoglobin 
content to be more accurate than the color index. 

The more carefully blood counts are made, the better the instruments 
used for hemoglobin estimation, 
the more evident are the individ- 
ual, daily, seasonal and racial va- 
riations. The regulation of the 
composition of the blood is won- 
derfully efficient. Although enor- 
mous amounts of water may pass 
through the vessels as in diabetes 
insipidus, of water and solids as in 
diabetes mellitus, of albumin and 
water as in cases of rapidly col- 
lecting ascites repeatedly tapped, 
yet the blood's composition in FlG> I5 ,_ Sodium biurate crystals from a tophus . 
any individual case varies within x 4 °°- 

comparatively narrow limits. 

The volume index or the quotient of the volume per cent, as determined 
with the hematocrit (considering a column of corpuscles of 50% as normal, 
see page 460) and the count per cent. (5,000,000 = 100%) promises to be 
of value. 44 The most important result of Capp's work in this field is that in 
pernicious anemia the color index never exceeds the volume index; that is, 
that there is no " supersaturation " of the corpuscles with hemoglobin but 
the high color index is due to an increase of the size of the cells alone. On the 
other hand, in secondary anemia the color index may fall below the volume 
index, while during regeneration the volume index may return to normal 
first. He has never seen any evidence of " acute dropsy" of the red cells. 

WHITE BLOOD-CELLS 

Granulations of Leucocytes. — By granules of leucocytes are meant 
in this section the minute bodies, usually spherical, with a size and staining 
character fairly constant for each granulation, which can be easily seen in 

44 Capps, Jour, of Med. Research, 1903, vol. v. 




492 CLINICAL DIAGNOSIS 

the fresh blood and which seem to be inclusions of the protoplasm of the 
cell. All protoplasm is slightly and indefinitely granular but leucocyte 
granules are definite inclusions of the cell protoplasm which are liberated 
as independent bodies when the cell breaks up. They would seem to be 
a specific product of the secretory activity of the cell, not the result of a 
degeneration of the protoplasm or an accumulation of the products of 
metabolism, or inclusions from phagocytosis. 

Ehrlich classified the granulations as : first the eosinophilic, acidophilic, 
or oxyphilic granules (a). These granules are coarse, about ip in diameter, 
spherical or slightly oval, quite uniform in size and color and so refractive 
that in the fresh specimen they appear black. From a mixture of stains 
these always take the acid ingredient. These granules have been found 
in the blood of every animal from the frog to man whose blood has been 
examined. They are of albuminous nature and contain iron. 

Amphophilic (/3). — These granules are described as varying in size 
from that of a granules to others much smaller. Some are said to take 
acid and others basic stains. In a mixture of eosin and indulin, however, 
all the /3 granules will take the latter. They are sometimes met with in 
the same cells as a granules, hence Ehrlich considers them a younger stage 
of these. This is said to be the only case in which 2 specific granulations 
are found in the same cell. They are met with in some white cells of the 
bone-marrow of man and of various animals (rabbit and guinea-pig) and 
in the peripheral blood of patients with certain anemias. They may, in 
leukemia for instance, explain the variations in the size and tint of the 
granules of some of the eosinophile cells. 

Basophilic and Mastzell Granules (7). — In the connective tissue and 
blood of all animals and of man are cells which contain large basophile 
granules. Those found in the tissues are the true Mastzellen. These stain 
best with dahlia, taking a metachromatic rather than a strictly basophilic 
tone resembling the tint of mucin, of which, indeed, some claim that they 
are composed. This tint is best seen if polychrome methylene blue is used. 
These granules are spherical or oval in shape and vary considerably in 
size in the same cell. Cells containing somewhat similar granules are found 
in normal blood. These are increased in leukemia while in some cases of 
pleural exudate a*nd of gonorrhea these may be the predominating leuco- 
cytes of the pus. Even Ehrlich admitted that these leucocytes may not 
be related to the Mastzellen of the tissues. They seem to differ in origin 
while their granules do not stain exactly alike. What is more, the 7 gran- 
ules in the cells of abnormal bloods are not exactly like those present in 
the cells of normal blood and of bone-marrow. Those jn leukemia blood, 
e.g., are more soluble in aqueous solutions than are those of normal blood. 
It is, therefore, at least possible that we have to do with 3 or more different 
granulations, or with the same granulations at different stages of their 
development. 



THE BLOOD 493 

Basophile Granulations (5). — These granules were originally described 
by Ehrlich as occurring in the mononuclear cells, as not staining by dahlia, 
therefore not y granules, and as occurring especially in the cells of bone- 
marrow. Later he seemed to consider them not as true granules but as 
nodes of the reticulum of the protoplasm. 

Neutrophile Granules (e) . — These granules which are extremely fine and 
dust -like are seen in the mononuclear cells of the bone-marrow, a few in 
the mononuclear cells with indented nuclei of the blood stream, the endo- 
thelial leucocytes of later writers, but they fill the common polymorpho- 
nuclear cell of the blood. Stained with Ehrlich 's triple stain, devised as a 
specific stain for them, they take a lilac color, but they also take an acid 
stain and so are called the " fine oxyphilic granulation," in contradistinc- 
tion to the eosinophilic or " coarsely oxyphilic " cells. While a somewhat 
similar granulation occurs in the blood of cattle, swine and sheep (Hirsch- 
feld) granules of exactly this size, arrangement and color are found only 
in man and hence they are considered by Ehrlich to be specific for man. 

In addition to the above mentioned granules are others seen only in 
specimens stained with polychrome methylene blue mixtures in the mono- 
nuclear cells especially. The fact that these granules can be demonstrated 
is said by some to destroy Ehrlich 's sharp distinction between granular 
and nongranular cells (see below) . 

Neussers Perinuclear Granulation. — In certain specimens stained with 
Ehrlich triple stain are seen, in the mononuclear leucocytes especially, but 
sometimes in all forms of the leucocytes, blackish-green granules which 
always appear attached to the nucleus. They vary much in size. They 
have often a glistening or refractile appearance. Neusser considered them 
as characteristic of "the uric acid diathesis" but it has been shown 45 that they 
are in reality artefacts which can be produced by variations in the time of 
heating and of the stain, which with some mixtures may be produced at will 
and which bear no relation to the output of alloxuric bodies in the urine. 

The Granulation of Lymphocytes. — In well-spread specimens stained by 
the various modifications of the Romanowski stain (but not by Ehrlich 's 
stain) fine violet granules are seen in about % of the lymphocytes, especially 
those with a fairly wide protoplasm margin and also in some of the large 
mononuclear cells. They are not always spherical; their size is between 
that of the a and e; few or many may be present in one cell, yet as a rule, 
they are not too numerous to count. They are not found in cells in smears 
of lymph-glands or of marrow. By their discovery Michaelis and Wolff 46 
considered that they had broken down the sharp line of demarcation drawn 
by Ehrlich between the granular and the nongranular cells. Ehrlich replied 
that while it cannot be denied that these were granules yet they cannot 
be considered as forming a definite granulation in the sense in which he 

45 Futcher, Centralbl. f. innere MecL, 1899. 

46 Virch. Arch., Bd. 167, p. 151. 



PLATE I 

Cells of Normal Blood. 

i. Small lymphocyte; small mononuclear. 

2. Eosinophile leucocyte. 

3, 4. Large lymphocytes. 

5. Transitional mononuclear. 

6, 7. Polymorphonuclear neutrophile leucocytes. 

8. Mastzell. 

Cells Found in Splenomyelogenous Leukaemia. 

9, 11, 17. Neutrophile myelocytes. 

10. Dwarf polymorphonuclear neutrophile leucocyte. 

12, 13. Transitional cells between myelocytes and polymorphonuclear cells. 

14. Eosinophile leucocyte. 

15. Lymphocyte. 

16, 19, 20, 21. Large mononuclears. 

18. Dwarf polymorphonuclear eosinophile leucocyte. 

22. Polymorphonuclear eosinophile leucocyte. 

Erythrocytes in Chlorosis. 

23. Cells found at the height of the disease. These are the "doughnut" or "pessary' 

forms. 

24. Cells from the same case as 23 during convalescence. 

POIKILOCYTES IN PERNICIOUS An^MIA. 

* 25. Battle-door form. 

26. Sausage form. 

27. Microcyte. 

28. Megalocyte. 

Nucleated Red Blood Cells. 

29. Mature normoblast. 

30. Immature normoblast. 
31, 32. Intermediate forms. 

33. Megaloblast. 

34. Normoblast with nucleus showing fragmentation or incomplete mitosis. 

35. Fuchsinophilic normoblast. 

36. Leucocyte of the same size as 37 shown for comparison. 

37. Large nucleated red cell. 

38. Intermediate nucleated red (or megaloblast), the "reptilian form." 



PLATE I 






4Hfek 



.^- ■•■■■■ 






V v# 






CELLS OF NORMAL BLOOD. 



6 



V 





»*^a, 




- 

v?<, y 












SPLENO-MYELOGENOUS LEUK/EMIA. 



W \ 



w 









AT HEIGHT OF DISEASE. 

ERYTHROCYTES 
IN CHLOROSIS. 


SAME CASE DURING 
CONVALESCENCE. 


29 30 


m 



LEUCOCYTE FOR 
ALL STAINED WITH EHRLICH'S TRIPLE STAIN COMPARISON 

AND DRAWN TO SAME SCALE. 



POIKILOCYTES 
IN PERNICIOUS AN/GMIA. 



32 

m 







NUCLEATED RED BLOOD CELLS. 



F. S. Lockieood. 



THE BLOOD 495 

Large Mononuclears. — The cells of this group may vary in size 
from those as small as lymphocytes to others the largest cells in the blood. 
They have a large, oval, vesicular, eccentrically placed, faintly staining 
nucleus, which indeed may be overlooked, and abundant weakly baso- 
philic protoplasm with or without a few granules according to the stain 
used. The larger cells of this group make up about i% of the leucocytes 
of the normal adult blood (Ehrlich) but the entire large mononuclear 
group varies from 5.6 to 8.1% average 7.2% (Mitter 10.8%. ) In normal 
blood the large cells form a definite group of almost uniform size but in 
pathological conditions, especially leukemia, typhoid fever and malaria, 
this group may be represented by cells of all sizes from that of lymphocytes 
to large giant-cells. (Plate II. The group 9-20 contains many.) 

Those of this group with much notched nucleus, the so-called " wallet " 
or " saddle-bag " nucleus, were called by Ehrlich Transitional cells (Plate 
I, 5), since he at first considered them intermediate stages between large 
mononuclear and polymorphonuclear finely granular cells. Although he 
soon abandoned this opinion (see page 450) the name is still in use. These 
cells are the largest of all in the normal blood. The protoplasm stains 
quite deeply and often contains a few neutrophile granules. These cells 
average from 1 to 3% of the leucocytes of normal blood. They are but 
older forms of the large mononuclear group. 

Phagocytes of the Peripheral Blood. — Evans called attention to the occasional presence 
in the blood of large mononuclear leucocytes derived directly from the endothelial cells 
lining the walls of the capillaries in spleen, lymph glands and bone-marrow, which are 
definitely phagocytic in character. These he called endothelial phagocytes. He denied 
their presence in normal blood. Later Mcjunkins claimed that these not only were 
present in normal blood but that even 7% of the leucocytes normally are these. 

McJunkins Method for the Demonstration of a Phagocytic Mononuclear Cell in the 
Peripheral Blood. 48 — Three cubic centimeters of blood are added to 2 c.c. of a 3.8% 
sterile sodium citrate solution that contains 1 % by weight of a good commercial grade 
of lampblack. The citrate-lampblack liquid is previously shaken vigorously in a flask 
to secure as even a suspension as possible and the 2 c.c. are measured at once into 15 c.c. 
graduated centrifuge tube. To this the blood is added as quickly as possible. To obtain 
the blood the palmar surface of a finger tip is painted with tincture of iodin, wiped with 
95% alcohol, dried, punctured deeply with an automatic lance and the blood allowed to 
drip directly into the tube. This citrated blood is mixed thoroughly by striking the 
lower end of the centrifuge tube with the finger and is then immediately filtered through 
a single layer of freshly laundered muslin into a second centrifuge tube in order to remove 
the gross particles of lampblack. This second tube may be prepared in advance by auto- 
claving it with the cloth pressed into its upper portion in the form of a cone. The cloth 
is moistened with a 3.8% citrate solution before the citrated blood is poured onto it. It 
is not advisable to filter the suspension of lampblack before adding the blood because it 
tends to filter clear. 

The filtered citrated blood is centrifugalized at a moderate speed for 15 minutes 
and at a high speed for 5 minutes. The tube is removed from the centrifuge and the black 
layer of leucocytes on the surface of the corpuscles is carefully drawn into a hemocyto- 
meter pipet with large bore. Such a pipet may be kept in 80% alcohol and washed 

48 Arch, of Int. Med., Jan., 1918, xxi, p. 59. 



496 CLINICAL DIAGNOSIS 

before using with sterile citrate. The pipet is shaken for i minute, a wide rubber band 
stretched over its ends and it is then transferred to an incubator at 37. 5 C. where it 
lies for 1 hour in a horizontal position. During this time it is removed and shaken for 
1 minute at the end of 15, 30 and 45 minutes. 

At the end of the hour the pipet is taken from the incubator, shaken for 5 minutes 
and cover-glass preparations made in the usual way. To stain this smear it is covered 
with 2 drops of a polychrome blood stain for 1 minute, then 4 drops of distilled water 
are added and the diluted stain allowed to remain on it for 2 or 3 minutes. This is washed 
off with water and the stain differentiated for 10 seconds in 0.02% yellowish water- 
soluble eosin, washed with water, dried and mounted in colophoniumxylol. 

On examination, the cytoplasm of certain of the mononuclear cells is seen to contain 
many carbon particles while that of the lymphocytes and the majority of the polypho- 
cytes cells contains none. The smears are surprisingly free from extracellular carbon. 
If it were not for the carbon within these cells some would be mistaken for large lympho- 
cytes although most of them fall in the classes commonly known as transitional and large 
mononuclear leukocytes. 

The average diameter of the mononuclear phagocytic cells closely approximates 
that of the polymorphonuclear neutrophile but many are smaller than the neutrophils 
and approach the larger lymphocytes in size while a few are larger than any neutrophile. 
The cell outline is usually round, but irregularities may be produced when the smear 
is made, or pseudopodia may have been projected. 

The cytoplasm of this class of cells which pick up the carbon particles is character- 
istic. Considerable lampblack is taken up by all of these cells and many contain so much 
that the character of the cytoplasm cannot be made out. 

The zone of the cytoplasm is wide at some point, due to an indentation or eccentric 
position of the nucleus. In preparations stained in the usual way this is, next to its 
phagocytic properties, the most distinguishing feature of the cell. In many the cyto- 
plasm forms only a distinct band about the nucleus but it may be fully as wide as the 
nucleus itself. The cytoplasm stains a paler blue than that of the lymphocytes. It 
may be quite free of protoplasmic granules, but many contain granules quite similar 
to those of the neutrophiles except that they are more filamentous. They are like those 
of the neutrophils in their oxydase reaction, but are usually less distinct. Typical dis- 
crete " azur "granules, such as occur so commonly in lymphocytes, have not been observed. 

The nucleus is round, oval, horseshoe or saddle-back in shape, or presents a broken 
irregular contour. In a few cells of this type the nucleus is stellate or divided into 
separate chromatin masses. This morphology appear to develop during the hour of 
incubation, but it is not certain that an occasional cell of this class with a broken nucleus 
may nc t be found in the blood. The only important characteristic of the nucleus is that 
it is vsry rarely both regular in outline and centrally placed. It stains less heavily 
than the nucleus of the neutrophiles. 

Polymorphonuclear Neutrophiles (Plate I, 6, 7). — These cells which 
make up from 70 to 72% (Miller 64.2%) of the leucocytes of the adult 
and from 18 to 40% of the child's are about iom in diameter although in 
a well-spread smear, in which case they have flattened out upon the glass, 
they may seem about twice this size. The nucleus is characterized by its 
polymorphous nature and its deep homogeneous stain. It may be a strand 
variously bent, or 2 or more small fragments connected by fine filaments. 
The protoplasm takes a faint acid stain and is well filled with the neutro- 
phile granules. These cells are the ordinary pus-cells of inflammatory 
exudates. They sometimes contain glycogen. 



THE BLOOD 497 

Eosinophiles (Plate I, 2). — These cells are of the same size or perhaps 
a little larger than the finely granular leucocytes which they resemble in 
every way except that their protoplasm is often slightly more abundant 
and is filled with eosinophilic granules. These cells make up from 2 to 4% 
(Miller 2.8%) of the normal leucocyte count. 

Mastzellen (Plate I, 8). — This name is given, perhaps incorrectly, to 
any cell with basophilic granules. These cells are usually about the same 
size as the preceding but more often are smaller. Their nucleus is poly- 
morphous, very faintly staining and often trilobed. The protoplasm con- 
tains a variable number of granules of different sizes, yet for the most 
part as large as a granules, which form a band around the nucleus. These 
granules are not stained by the triple stain, hence one sees only the spaces 
which they occupy. (These are probably the reticulated or vacuolated cells 
of Uskow.) They stain best in thionin and are said to take a metachroma- 
tic tone. These cells make up about 0.5% (Miller 0.6%) of the total count. 

In addition to these the following leucocytes may be found in patho- 
logical conditions: 

Myelocytes (Plate I, 9, 11, 14, 17). — While any cell of the bone-marrow 
is, strictly speaking, a myelocyte, by this term is generally meant 1 with 
granular protoplasm and a round nucleus. Some are neutrophilic, some 
eosinophilic, while some claim to have seen true basophilic myelocytes. 
Neutrophile Myelocytes. — The size of thesecells varies from thatof the large 
mononuclears to that of red corpuscles. The largest and smallest neutro- 
phile myelocytes are found in the blood only in myelogenous leukemia, but 
a few neutrophilic myelocytes the size of the granular leucocytes may be 
found in any condition with a high leucocyte count. The characteristic 
point is the shape of their nucleus which is either perfectly round, oval, 
indented or kidney-shaped, but never polymorphous or pycnotic; if it 
were, the cell would count as an ordinary leucocyte. It is usually centrally 
placed. It is impossible to draw a sharp line between a myelocyte and a 
polymorphonuclear cell (Plate I, 12, 13) since every possible gradation 
occurs, but as myelocytes we count all granulated cells with round, oval, 
or kidney-shaped faintly staining nuclei, providing the nucleus apparently 
occupies at least % of the cell. A cell with nucleus relatively smaller, more 
compact, more distorted and more diffusely stained ranks as a leucocyte. 
The nucleus of some leucocytes is round or oval but it also is relatively 
small (occupying only about a quarter of the diameter of the cell) and stains 
deeply and diffusely. The chances are that could we get a side view of 
these nuclei we would find them polymorphous. For the question of the 
motility of myelocytes, and most now agree they are ameboid, see the 
writings of Wolff. 49 

Some myelocytes are full of granules while others have but few and 
these are scattered. The very large forms are met with in the bone-marrow 

49 Deut. med. Wochenschr., March 5, 1903. 
32 



498 CLINICAL DIAGNOSIS 

and in well-made specimens of blood of cases of myelogenous leukemia, 
but as a rule one sees only a large faint nucleus surrounded by granules 
free in the plasma. 

Eosinophile Myelocytes (Plate I, 14). — The coarsely granular myelocytes 
are the exact analogue of the preceding and occur under much the same 
conditions, but less often and in much smaller numbers. They are found 
especially in splenomyelogenous leukemia and in anemia pseudolymphatica 
infantum. 

Small Neutrophiles: Pseudolymphocytes . — The small neutrophile leuco- 
cytes have a round, intensely staining nucleus and a narrow margin of 
protoplasm full of neutrophile granules. Their size is about that of a 
lymphocyte. They are rare, occurring especially in pleuritic exudates and 
are supposed to rise from fragmentation of the polymorphonuclear cells. 

Irritation Forms. — The so-called "irritation forms " vary in size from 
a lymphocyte to a large mononuclear, but the majority are small. Their 
nucleus is round, of a bluish-green color (Ehrlich stain), often eccentric 
and has no chromatin net-work. Their protoplasm stains an intense rich 
brown and has no granules. Turk says that they occur under the same 
conditions and have the same significance as myelocytes. 

Differential Counting. — In making a differential count one first must 
agree on certain rules of classification which are more definite than our 
actual knowledge would justify. Since we know so little of the relationship 
between the various forms of leucocytes, their ages, their origin and of their 
function, the only classification possible is a purely morphological one. 
We have rules which should be followed mechanically if our results are to 
be at all comparable. We separate first granular and non-granular cells. 
This in a well-stained specimen should be easy. There is no difficulty in 
separating the coarsely granular and the finely granular cells. Whether 
or not we are justified in separating the coarsely granular cells into two 
groups, the eosinophils and Mastzellen, using specimens stained with 
methylene blue, is an open question. We do, however, make a separate 
classification for the Mastzellen. 

In normal blood it is very easy to distinguish between the small and the 
large mononuclears but in some pathological conditions in which large 
lymphocytes and small endothelial leucocytes are present we must fall 
back on the unsatisfactory rule that lymphocytes are smaller and large 
mononuclears are larger than a polymorphonuclear neutrophile leucocyte. 
For normal blood this classification is satisfactory but in pathological 
conditions many objections arise. While the lymphocytes seem to form a 
fairly distinct class although some are large (Plate I; 3, 4, 15, 20), the group 
of endothelial leucocytes contains large, medium and small forms and any 
line based on size which divides this group is arbitrary and increases the 
number of the lymphocytes by cells which do not belong there and dimin- 
ishes a group which should not be divided. This is best seen in typhoid 



THE BLOOD 499 

fever and malaria, diseases in which the group of endothelial leucocytes is 
increased and this group then includes large and small forms. In the so- 
called lymphatic leukemia also the small mononuclear cells are certainly 
not all lymphocytes. 

Neither can we separate groups on the basis of an indentation of the 
nucleus. Ehrlich's name ' ' transitional ' ' is still used although be abandoned 
it as soon as he discovered that the granular myelocyte and not the large 
mononuclears with indented nuclei, the so-called " transitionals," were the 
young forms of polymorphonuclear granular cells. 

The line between myelocyte and leucocyte is very hard to draw (see 
page 498). 

In leukemia it is very hard to draw a line between those large mono- 
nuclears (Plate I, 16, 19, 21) with deep staining protoplasm and the granu- 
lar myelocytes and perhaps no such line exists. Yet in well stained speci- 
mens one is in doubt concerning but few cells. 

One group of cells is confusing, those represented merely by a faint mass 
of stain. Such cells should be counted as " undetermined cells " for only 
in that way will the percentage of those groups which are more resistant 
be fairly correct. These undetermined cells are almost all non-granular 
small mononuclears. 

There should theoretically be little difficulty in differentiating eosino- 
phils and neutrophiles and yet even in well-stained specimens the question 
may be so hard that it leads one to doubt the specificity of granules. We 
believe that now most observers no longer use Ehrlich's stain and so do 
not look for the characteristic lilac tint of the neutrophile granules, but 
consider all finely granular cells as neutrophiles and all coarsely granular 
ones as eosinophiles. 

Ehrlich's neutrophilic granulation has not gained the clinical import- 
ance which he anticipated. It certainly would be fairer to use the term 
" finely granular cells " or " fine acidophilic," in case other stains are used 
and to reserve the term neutrophilic for cells stained with his triple stain, 
for that is the only successful specific stain we have for those granules. 

For differential counting the best of specimens are none too good and 
far too much time is spent over smears which should be thrown away. We 
still prefer specimens stained with Ehrlich's triple stain but use those 
stained with Hasting 's stain. Nucleated reds are counted at the same time 
as the white cells and calculated as " number per cu.cm." 

The list of types recognized is, therefore, the following: Small mono- 
nuclear (s. m.) large mononuclear and transitional (l.m. & tr.) ; polymor- 
phonuclear neutrophile (pmn. e) ; polymorphonuclear eosinophile (pmn. a) ; 
Mastzell (Mastz.); neutrophile myelocyte (myeloc. e); eosinophile myelo- 
cyte (myelloc a). Nucleated reds; normoblasts (normobl.), intermediates 
(intermed.) and megaloblasts (megalobl.). 

To make a differential count a mechanical stage should be used and at 



500 CLINICAL DIAGNOSIS 

least 500 leucocytes counted. Some keep count with a pencil and paper, 
1 column for each group ; others use a slide-box divided into compartments 
by slides, into which he drops beans, one bean for a cell. Since one can 
start with exactly 500 beans the mathematics of this calculation is easy. 

BONE-MARROW 

The careful study of the bone-marrow should be required of each student. In it 
are found normally practically every cell which ever occurs in the blood and a complete 
series of transitional forms between the apparently related groups. 

The study of marrow while fresh is especially valuable, especially that of the ribs 
of young babies or premature infants, but fragments of ribs removed for empyema and 
aa autopsy will suffice if fresh enough. A small piece of rib is squeezed in a pair of forceps 
and a drop of the exuding marrow picked up on a cover-glass and at once pressed down 
onto a slide. Very rapid work is necessary, since the drop dries very rapidly. It is 
surprising how quickly some of the interesting mononuclear forms, the " young " cells, 
disintegrate. The large myelocytes also soon disappear and in leukemia the marrow 
smear may soon show only a confused mass of nuclei in a cloud of free granules. For 
stained specimens, the stroke method is best; that is, the marrow is smeared in lines on 
the cover-glass by drawing this across the end of the bone. The smear is allowed to 
dry in the air and is then fixed and stained just as are blood smears. If the marrow is 
fatty the smears do not stain well. If they are to be fixed by heat they should be over- 
heated, best on the copper plate, smear side up, at the spheroidal point (that is, the point 
at which the drop of water does not boil but merely rolls off the plate) for 45 or more 
seconds. Such specimens will have some good areas for study, especially at the edges 
of the thick portions, and a few such fields are all that is desired. Thin well-spread 
specimens are usually failures. Better a specimen generally too thick but with some 
thinner areas. 

The bone-marrow of certain vertebrates would seem to be made up of a mosaic of 
little separate masses of different tissues, each concerned with the production of 1 type 
of cell. These islands cannot be demonstrated in human marrow and yet this would 
seem to contain quite distinct tissues more diffusely arranged. In some places will 
be found nests of nucleated reds in enormous numbers; in other places nests of leuco- 
cytes, myelocytes and of intermediate forms. Different parts of the same rib vary much, 
as we have found to be the case in infant marrow. Since the marrow varies so in different 
bones and in different parts of the same bone, it is impossible from a limited search to 
say what is the general medullary condition of a given case (Grawitz) . This may explain 
the lack of evident relation between a marrow and a blood picture. 

NUCLEATED RED— Blood-Cells. By the term "erythroblast" most writers mean 
any nucleated red blood-cell. A few, however, use it of a hypothetical colorless ancestral 
form of these. A better term for these would be " hemoblast." 

(1) Normoblasts. — (a) Howell's Mature Nucleated Reds (Plate I, 29). — These cells 
have the color of the non-nucleated red blood-corpuscles. Their nucleus is 3jU or slightly 
less in diameter, is sharply denned and is pycnotic; that is, it is dense, homogeneous, 
structureless (triple stain) , takes a dense uniform blackish-green color, has no chromatin 
net- work and is often vacuolated, hence often has a bright spot in the fresh and an 
unstained area in the stained specimen. They often present amitotic figures, i.e., 
rosette forms of 2 to 4 or even 12 fragments connected by strands of chromatin (see Plate 
I, 34; Fig. 113, c). The nucleus is often surrounded by a clear zone. These nuclei are 
so characteristic that they may be recognized even if not surrounded by protoplasm, 
which is often the case since many are found resting upon a margin of the red cell or 
even at some distance from it. This is due, said Ehrlich, to the centrifugal force which 
throws the heavy nucleus out of its cell when the specimen is made. Yet this can- 
not explain all the free nuclei which are seen in specimens made in various ways and 



THE BLOOD 501 

in cut sections as well. Pappenheim and Israel claim that in leukemia especially such 
free nuclei result from the degeneration of their surrounding protoplasm. 

(b) Howell's Immature Nucleated Reds (Plate I, 30; Fig. 113, a). — The immature 
nucleated reds are a little larger than an ordinary red blood-cell, their color is perhaps 
a little paler and their nucleus slightly larger, than that of the mature form. They 
have definite chromatin fibers arranged radially (in leucocytes they form a meshwork) 
while mitotic figures are not rare. The division of these cells is rapid, requiring but 
15 minutes. In the bone-marrow this is the dominant red cell. Between these 2 cells, 
the mature and the immature, one finds all intermediate stages. 

These 2, and all intermediate, forms of nucleated red cells are generally called normo- 
blasts, although the larger immature forms are by some classed as intermediates. They 
are the precursors of the non-nucleated red blood-cell but do not reach the circulation 
of a normal adult except as an anomaly. Their appearance in the general circulation 
indicates an increased activity of the bone-marrow. Many are met with in the blood 
in pernicious anemia, more in splenomyelogenous leukemia and some in post-hemor- 
rhagic anemias. They appear in the blood of a child more readily than in that of an adult. 

These cells normally are " orthochromatic "; that is, they stain like the ordinary 
non-nucleated reds {i.e., are oxyphilic), but some are fuchsinophilic while others are 
basophilic (polychromatophilic) . 

Blood crisis is the name given by v. Noorden to the appearance in the circulation 
of enormous numbers of nucleated reds and an increase in the leucocyte count, which 
occurs usually during the convalescence for an anemia. They remain in the circulation 
for a few days after which there usually is a sudden rise of the red blood- count. The 
nucleated reds then slowly disappear and the increase in the count is much less rapid 
until perhaps another crisis occurs. V. Noorden reported cases with gains in the counts 
of half a million cells in 4 days. Blood crises are supposed to indicate a temporary 
increase in the activity of the bone-marrow. They are, however, not always a sign of 
improvement (see page 608) but always indicate a struggle which may prove unavailing. 

Intermediate Red Blood-Cells (Plate I, 31). — By the term intermediate nucle- 
ated red blood-cell is meant one which is not quite large enough to be called a megalo- 
blast and yet which is definitely larger than a normoblast. Some are large cells with the 
nucleus of an immature red; others are smaller cells with a reticular nucleus larger 
than that of a normoblast. If one systematically measures all nucleated reds in a spe- 
cimen he will find very few of these cells unless one includes the immature nucleated 
reds of Howell (Fig. 113, b). 

Megaloblasts. — In the marrow are always found nucleated reds (Fig. 113, f) 
which are from 2 to 4 times the size of an ordinary red blood-cell. They are round or 
oval, their protoplasm is abundant and often polychromatophilic and their nucleus is 
large plump, round or oval, centrally placed and conspicuous, while in stained specimens 
a good chromatin net-work is demonstrable. These cells are quite similar to the large 
nucleated reds seen in primary anemias and called megaloblasts. 

To understand a writer's report of a blood case it is necessary to know his definition 
of a megaloblast. Opinions vary much. Some demand that the cell be large, others 
that it have a large nucleus. Our rule is that both of these specifications must be ful- 
filled, and, for reasons to be given later we ask that the size of the nucleus shall be at 
least that of an ordinary non-nucleated red (7.5/x). Pappenheim and others consider 
the megaloblast a different type of cell from the normoblast and say that it may be recog- 
nized by certain fine points in the nucleus even though the cell be as small as a normoblast. 

The megaloblast of the circulating blood in pernicious anemia differs somewhat from 
the above mentioned large nucleated red cell of the bone-marrow. They are about 
equal in size but the normal bone-marrow megaloblast usually has a round nucleus with 
a very distinct margin and a definite chromatin net-work, while the nucleus of the 
megaloblast in the blood in pernicious anemia often is more oval, much less distinct, 



502 CLINICAL DIAGNOSIS 

stains much fainter and has less definite nuclear membrane and chromatin structure. 
But these differences are slight and inconstant. 

The significance of megaloblasts in the blood has been the subject of much contro- 
versy. Ehrlich claimed that they are never found in the normal bone-marrow of- an adult, 
but rather are the product of a megaloblastic degeneration of this tissue due to a toxine 
which brings about a reversion of the marrow to the embryonic (others say " to the 
amphibian ") condition. Ehrlich well said that any attempt to break down the distinc- 
tion between normoblasts and megaloblasts fails from the fact that in pernicious anemia 
the entire blood picture is megaloblastic. (We are inclined to think that the expression 
" reversion to the amphibian type of blood " is much too often used. The only amphi- 
bian we have studied whose blood resembles that of pernicious anemia is the batracho- 
seps, and one would hardly call an hemoglobinemia a " reversion," although that is 
the blood condition in some worms). While a " megolablastic degeneration " may 
explain the large numbers of megaloblasts in the marrow of cases with pernicious anemia 
and in some other conditions, we fail to find any recent observer who has not found 
them in all normal marrows. In our studies of bone-marrow we have measured many 
nucleated reds and have found the predominant cell the immature normoblast, with a 
nucleus between 3 and 4/x in diameter. The next most common cell, about 15% of all 
nucleated reds, is a megaloblast, with a nucleus of 7/x or over in diameter, or 7/x in 1 axis 
if oval. Between these 2 groups of cells occur every intermediate size, and yet their 
number is not as great as that of the large cells. 

Bunting has shown that in certain animals the marrow contains islands of nucleated 
reds in the center of which are megaloblasts surrounded by zones of cells of different 
size until one reaches to the periphery where normoblasts are found. The activity of 
these zones produces cells of the same size as well as those for the layer external to it. 
Ordinarily the cells for the circulation are produced at the periphery of these islands 
but if the islands are over-taxed or affected by certain poisons the successive layers are 
in part stripped off and larger cells of the interior may send cells into the general circula- 
tion. This could be a very satisfactory explanation of the megolablastic character of 
the blood in pernicious anemia and in the bothriocephalus anemia. It might also explain 
the blood crises, etc. 

Karyokinesis of these large cells occurs in the peripheral blood in severe anemias, 
especially as a terminal phenomenon. 

Thayer has seen definite ameboid movements in a megaloblast. 

Microblasts. — By microblast is meant a nucleated red under 6jjl in diameter with 
a small pycnotic nucleus. They occur in the circulation in severe traumatic anemias, 
leukemia, etc. Some appear to be perfect cells and these may be the forerunners of 
microcytes, but others would seem to be pinched off from larger cells. 

The fate of the nucleus of the red cell is still the subject of much discussion. Two 
views have been held: (1) that the mature normoblast extrudes its nucleus (Rindfleisch, 
Howell, e.g.), and (2) that it disappears within the cell by karyorrhexis and ka^olysis 
(Kolliker, Neumann, e.g.) . Those who hold the latter opinion admit that some nucleated 
red cells may be seen to extrude their nuclei, but consider it pathological. Other writers 
believe that all of these methods are possible; Ehrlich, for instance, thought that the 
normoblastic nucleus is extruded but the macroblastic is absorbed. In favor of the extru- 
sion theory is the fact that most non-nucleated red cells are flat, disk-shaped and even 
biconcave. But some are even spherical, especially in the embryo, a point emphasized 
by those holding the theory of absorption. The " degenerations " or nuclear fragments 
described by Vaughan (page 449) and by Cabot (page 477) would suggest karyorrhexis. 
The free nuclei so often seen in stained specimens may have been thrown out of the nor- 
moblasts by the centrifugal force caused by the sudden motion of the cells when the 
specimen is made and the reason why the nucleus of the megaloblast remains in the 
cell may be that its specific gravity is nearer that of the protoplasm. 



THE BLOOD 503 

The changes in the nuclei are important. Bya " pycnotic " nucleus is meant I 
diminished in size, dense,homogeneous, sharply defined, sometimes vacuolated and with- 
out any evident chromatin net-work, all of which suggest a solution of the 
chromatin in the nuclear fluid. Pycnosis may be a preliminary step of karyolysis 
or absorption although in the latter case the nucleus as a rule becomes fainter till it 
cannot be distinguished from the surrounding protoplasm. Pycnosis may precede 
karyorrhexis or fragmentation of the nucleus, which fragments may then disappear by 
karyolysis. The normoblastic nucleus may by amitosis divide into polymorphous forms 
with 2 or even 12 fragments (Plate I, 34) of equal or unequal size and usually united by 
a filament. In 1 of our cases during a blood crisis 55% of the erythroblasts were of this 
description. Another method of nuclear destruction is suggested by cells containing 
only a few chromatin strains and masses as though nuclear membrane and fluid dis- 
appear first. 

A point evident in bone -marrow specimens is the varying depth of the hemoglobin 
tint of the red corpuscles, a variation much greater than is seen in the blood. This 
might suggest that the formation of hemoglobin is a gradual intracellular process. This 
is a far-reaching problem not only in cytology but also in diagnosis (see page 592). The 
question is of particular interest in the study of the anemias. Are these pale cells in the 
circulation permanently pale or are they only immature and later develop more hemo- 
globin? When the color-index falls is it because cells are losing hemoglobin or because 
new light-weight cells are replacing heavier ones? As the case improves do cells develop 
more hemoglobin? That is, is the cell like a coal cart which carries different loads at 
different times? Arguments from comparative anatomy are not satisfactory, yet the 
red cells of some of the lower vertebrates would seem to complete their development in 
the circulation. In the mammals an imperfect cell is said to be incapable of further 
development. Finally, Gaule and his pupils believe in a hemoglobin " store " in the 
body, which hemoglobin in case of need is carried into the circulation in new corpuscles 
and returned when the extra cells are withdrawn. In adult man the evidence would 
tend to show that a red cell is the product of erythroblastic tissue not of hemoglobin 
free tissue, that each cell in the circulation whether complete or incomplete is finished 
so far as that cell is concerned ; that the formation of hemoglobin is a slow process, much 
slower than is the proliferation of new cells ; and that the bone-marrow, when the demand 
for new cells is heavy, uses its available store of pigment to make light-weight cells and 
later replaces them with cells of normal weight. This is reasonable since red cells are 
functionally valuable in proportion to their surface, not their weight. 

Origin of Red Blood-cells. — That the ordinary non-nucleated red blood-cells 
come from nucleated reds is now doubted by few. In the embryos and possibly later 
under conditions of increased hematopoiesis there is also the intracellular differentiation 
of erythrocytes within the hemogenic polykaryocytes' (see page 506 ). 50 Up to the end 
of the fourth week of embryonic life all of the blood-cells are nucleated. From that time 
on the number of the non-nucleated cells increases so that at the third month only about 
% to % are nucleated. At the fifth month nucleated reds are still numerous but at birth 
it is seldom that one is found in the blood. 

In the earliest embryonic life the blood-vessels are formed from solid cords of cells 
the peripheral cells of which become the endothelial lining of the vessel wall, the internal 
cells the corpuscles. This process may occur in almost any part of the developing organ- 
ism and may also in the adult wherever new blood-vessels are formed. In the embryo 
also many mitoses are found in the nucleated reds of the circulating blood. 

Before the third month the liver has become the chief seat of blood formation, after 
the fifth month the spleen and the lymph-glands take up the task but finally the marrow 
becomes the chief blood -building organ. In the child the marrow of the whole skeleton 

50 Jordan Am. J. of Anat., 1918, Vol. 24, p. 225. 



504 CLINICAL DIAGNOSIS 

has this function but at about puberty and during adult life only the ribs and some of 
the flat bones. Howell considers that callus, for instance that following a fracture, 
may in the adult for a while furnish centers for hematogenesis. In the adult it would 
seem as if the spleen could resume this function in leukemia and anemia. 

Removal of the spleen causes very little anemia, but about a month later the small 
mononuclears begin to increase and this continues for months resulting sometimes in a 
blood picture which strongly suggests leukemia. About 12 months after the splenec- 
tomy an eosinophilia of even extreme degree may develop. These phenomena are now 
explained as evidence of the vicarious activity for the spleen, first by the lymph-glands 
and then by the bone-marrow. In health one of the functions of the spleen seems to be 
to remove old and injured red cells and leucocytes from the circulation and the acute 
spleen tumor in some conditions may be due to the great number of leucocytes ingested 
(spodogenic splenic tumor). 

In the embryo the blood-cells at first are without hemoglobin. At this time also 
there are no true leucocytes and none appear until after the formation of hemoglobin- 
containing cells is quite active. The embryologists have shown that in disease of the 
embryo before the appearance of leucocytes the red blood-cells are ameboid and perhaps 
phagocytic, which is interesting since in certain blood diseases of the adult one gets 
hints at least of these 2 functions. 

The view is still held by some that in some forms of acute lymphatic leukemia the 
abnormal white cells are " red cells " without hem'oglobin, and the converse of this 
also is held by some to be true, that is, that in severe anemias some of the large lympho- 
cytes develop to megaloblasts instead of to small lymphocytes (Pappenheim) . 

Howell was the first to demonstrate in the cat the " ancestral red corpuscles " which 
resemble the red blood-cells of reptiles since they are large, oval, semifluid red cells, 
with a deeply stained oval nucleus. These cells were later described by Engel as ' ' metro- 
cytes of the second generation," those of the first generation having a large chromatin - 
rich nucleus. Such cells later never, normally at least, reach the circulation and Engel 
thinks they are no longer formed. 

For the study of the young red cells the blood of embryo mice is to be especially 
recommended. Here a great variety of changes in the nucleus and of granulation of the 
protoplasm may be demonstrated. 

White Blood-Cells. — In the marrow one finds: A. Granular Cells. I. Neutro- 
philes. Neutrophile myelocytes. 

1. Typical myelocytes are cells from 12 to 15^ in diameter, with a large round 
nucleus and protoplasm scanty and finely granular which forms often a thin rim around 
the nucleus. These cells are by far the most numerous in the marrow but on account 
of their size seem even more so than is the case. The nucleus is often hard to make out. 
In the bone-marrow may be seen also the much larger beautiful myelocytes with faint 
nuclei, " Cornil's marrow cell," which however are not often seen intact since they 
disintegrate so readily. These appear in the blood only in leukemia. In the fresh mar- 
row some of the myelocytes have a very small, dense, round nucleus, which we think 
is due to post-mortem loss of nuclear fluid. All transitions from typical myelocytes 
with round nuclei to typical leucocytes are present on the one side (these transitional 
forms are called " Metamyelocytes "). and all transitions from typical myelocytes full 
of granules to large mononuclears with clear protoplasm (myeloblasts) on the other. 
The transitional forms between myeloblasts and myelocytes are called " promyelo- 
cytes." 

2. Cells similar to the above, but much smaller, their nucleus indented, or slightly 
polymorphous and staining faintly, are called transitional cells (between 1 and 3). 

3. Typical leucocytes. 

II. Eosinophiles. — These always are relatively few in number. 

I. Eosinophile myelocytes. Large cells with pale nuclei, scanty protoplasm filled 



THE BLOOD 505 

with eosinophile granules, otherwise similar to the above mentioned neutrophile cells. 

2. Similar to the above, but smaller; the nucleus indented or slightly polymorphous. 
All gradations are seen from this to typical — ■ 

3. Eosinophile leucocytes. 
III. Basophiles. 

1. Mastzellen, which, however, are rather rare. The nuclei of these cells may have 
a variety of shapes, yet are usually polymorphous (see page 498) . Mononuclear Mastzel- 
len occur (Engel). These are, at least, hard to recognize, since the young (?) a and e gran- 
ules are quite basophilic. 

2. Cells varying in size but the most of them small and containing violet colored 
granules with basic stains. These, although they contain basophile granules, are not 
typical Mastzellen. In this connection it should be mentioned that the fi granules may 
occur in the eosinophile cells or by themselves. 

3. Polymorphous cells, with fine basophile granules. These may be neutrophile 
cells stained by the " tricky " methylene blue. 

B. Non-Granular Cells. — 1. Lymphocytes. These have the size of red blood- 
cells, a narrow rim of protoplasm and a nucleus rich in chromatin. These cells are the 
second most numerous cells of the marrow and often resemble naked nuclei. 

Among these are the " protoleucocytes " of Osier; solid-looking lymphoid elements 
from 2.5 to 5ju in diameter, which resemble free nuclei; some have a rim of protoplasm. 
From these " erythroblasts (?) " develop. 

2. Medium-sized lymphocytes with more protoplasm and a smaller and often 
eccentric nucleus. 

3. Very large cells with general character of lymphocytes which appear in the blood 
in some acute leukemias, but never normally. These are the " Large lymphocyte " 
(Ehrlich, Frankel, Pappenheim); Grawitz's " unripe cell," Wolff's " indifferent lymphoid 
cell," Naegeli's " myeloblast," Troje's " marrow-cell." Their nucleus stains faintly, 
is seldom lobulated, is very pale and poor in chromatin and their protoplasm is faintly 
basophilic. 

Also to be mentioned are cells, common enough, which in the fresh marrow resemble 
normoblasts (immature), except they have no hemoglobin. They are from 9 to i2f± in 
diameter. Their nucleus is that of Howell's immature red (see page 502), and they have 
a hyaline protoplasm. These are the " erythroblasts " of Osier, Lowit, and Howell. 

Lowit described as " leucoblasts " cells with relatively large nuclei which contain 
1 or 2 chromatin masses which are sometimes irregular in shape and from which a system 
of delicate lines and bands radiate to the distinctly doubly contoured nuclear membrane 
which has on its inner surface projections connected with the infranuclear net- work. 

Ehrlich believed the true lymphocyte came from the lymph-glands ; others say only 
from the bone-marrow. Some have tried to distinguish morphologically those from the 
marrow from those from glands (Rubenstein) . The question now is, " Do any come from 
lymph-glands? " thus admitting that typical " lymphocytes " are an important constitu- 
ent cell of the marrow. Michaelis and Wolff tried to differentiate these cells on the basis 
of their future history, the lymphocytes from lymph-glands remaining such, while the 
" lymphoid " cells of the marrow were capable of further development to a granular 
cell. But this " capability " would be hard to determine in the case of the individual 
dead cell now before us, although these writers did also describe slight differences in 
their staining reactions. And yet a distinction between lymphocytes and lymphoid cells 
is probably quite just (see page 469). 

Many workers considered these lymphoid cells as young forms and named them 
" protoleucocytes " (Osier), etc., considering that from them develop the colorless cells 
which correspond to the leucoblast and the erythroblast of Lowit and Howell, from 
which develop the whole series of the red and the white cells. The lymphocyte with 
diffusely staining notched nucleus (Rieder's cell) is probably an old form of lymphocyte. 



506 CLINICAL DIAGNOSIS 

The small mononuclears of the bone-marrow with round vesicular nucleus, delicate 
chromatin net-work and rather broad band of basophilic protoplasm with smooth mar- 
gin, are young cells which resemble those of the normal blood only in appearance but 
themselves do not belong there. The lymphocytosis of young babies, the most rational 
explanation of which is an overproduction of leucocytes to fulfill a function which did 
not formerly exist, would suggest that young lymphocytes are lymphoid cells. 

In the marrow are cells which never reach the normal circulation. Among these 
are young (also old?) leucocytes and red blood-cells and some say the undifferentiated 
cells which are the ancestors of both series, although we follow Bizzozero that none of 
the ancestors of red cells are without hemoglobin. The trouble is that there is such a 
variety of cells found in the marrow that one may find evidence in favor of any ancestral 
tree he wishes to draw. Formerly the series were traced back to small cells but now the 
cells considered young are very large, with large, faintly staining nuclei and protoplasm 
which very quickly goes to pieces, hence they are seldom seen. Probably too much 
stress has been laid on the shape of the nucleus as evidence of age. It may be that 
irregularity in the nucleus is merely an expression of function. 

Most will agree that there is in the marrow a large group of indifferent cells which 
may develop in some direction or other, possibly to red or white series as necessity 
demands. The only question is, " Which are these cells? " 

The development of cells is more by " steps " than by a gradual transition and those 
of each step are able to produce others of their kind as well as some of the succeeding 
generation. The picture as usually drawn is very complicated, since the line of descent 
of these cells is not single but several ancestors may produce the same forms so that to 
trace the description backward is more like following a stream toward its source. It is 
pictured as a single river at its mouth, but as we go toward its source many tributaries 
are found which contribute to its volume. Thus Pappenheim says normoblasts come 
from small lymphocytes, megaloblasts from large lymphocytes, and considers the poly- 
chromatophilic group as evidence of the transformation from a basophilic lymphocyte 
to a red cell; subsequent workers trace normoblasts from megaloblasts. Normoblasts cer- 
tainly can produce normoblasts and megaloblasts, megaloblasts. Again the granulation, 
may appear in cells with nuclei at various stages of deformity as if the changes in the 
nucleus from round to polymorphous bore little parallelism to the development of granules. 

We cannot here take up the question of the origin in other organs of leucocytes and 
perhaps of red cells. The above remarks are not intended as a resume of the subject, 
but an answer to many of the questions asked by students while studying the smears of 
bone-marrow. We will merely mention Nothnagel's case of general osteosclerosis with 
the entire marrow practically functionless, yet with a normal count of neutrophiles ; 
also the presence of mononuclear granular cells in areas of inflammatory infiltration. 

4. Pigmented cells, often absent. 

5. Giant-cells. 51 — (a) Megakaryocytes with 1 large irregularly coiled nucleus. These 
" giant-cells with budding nuclei " are considered as the parent cells of the blood plate- 
lets. 52 Jordan believed that the megakaryocytes (polymorphokaryocytes) are hyper- 
trophied hemoblasts; that their basket nucleus undergoes direct division producing 
polykaryocytes which are hemogenic giant cells. These are essentially multiple hemo- 
blasts comparable to the blood-islands of the yolk-sac and under certain conditions, 
apparently those of increased hematopoietic activity, may differentiate erythrocytes 
intracellularly. Jordan suggests that these cells represent an incidental phase of intense 
hematopoiesis ; he grants that certain specialized megakaryocytes may serve as sources 
of origin of blood platelets (according to the conclusion of Wright). 

Jordan does not believe that these cells are phagocytic. 

51 Jordan Am. J. of Anat., 1918, xxiv, p. 225. 

52 Wright Jour. Morph., xxi, p. 265. 



THE BLOOD 507 

(b) Giant phagocytes, always mononuclear and resembling large lymphocytes or 
endothelial leucocytes, their giant size due to their swollen condition from the large 
number of ingested erythrocytes. These cells are seldom found in the stained specimen 
although one does find masses of detritus which may represent them. These cells may 
enter the circulation in cases with marked leucocytosis and are filtered out in the lung 
(see Plate II, ii). 

(c) Multinuclear osteoclasts which are not related to hemoblastic polykaryocytes. 
These also are polynuclear. 

Many cells are seen, especially in fresh specimens, which contain very interesting 
degenerations and inclusions. Some large mononuclear cells contain large globules or 
droplets about 3^1 in diameter which are rather uniform in size and have the yellowish 
shimmer of the myelin droplets of the sputum. Some cells are filled with very large 
granules which have the color and retractility of a granules (see Fig. 115, e). Howell 
found many such cells in the marrow of the cat and thought that they must play an im- 
portant part in metabolic changes in the marrow. In other cells one sees globules of 
fluid which give them a vacuolated appearance. In some cells large " dropsical " pro- 
jections both of protoplasm and of the nucleus are seen. 

It is much easier to trace the degenerations of the leucocytes than of the red cells. 
In normal blood practically all the leucocytes are normal, but when there is a leucocytosis 
and especially in the leukemias one finds many cells with definite degenerations. The 
lymphocytes may be almost devoid of protoplasm, their nucleus is small, pycnotic and 
indented, or even polymorphous (Rieder's cells) (Plate II, 17). 

The nuclei of the polymorphonuclear granular leucocytes are very pycnotic and 
fragmented although probably all the fragments are corrected by a chromatin thread. 
For an interesting classification of the neutrophile leucocytes based on the number of 
nuclear fragments, and the clinical use to which such a classification may be put, see the 
publications of Arneith. 53 

The large pale nuclei without protoplasm seen in the marrow are said to be the nuclei 
of very sensitive young cells the protoplasm of which is destroyed in the preparation of 
the specimen, but similar appearances in lymphatic leukemia suggest very strongly that 
these may be only degenerated lymphocytes. 

Late in leukemia it would seem (said Ehrlich) as if the ability of the marrow to 
develop neutrophile granules might be lost, and clear cells are found which resemble 
the granular cells in every way except that their protoplasm is clear. 

Do the blood leucocytes have the ephemeral history which some ascribe to them? 
Winternitz (quoted from Grawitz) estimated that in the dog the lymph supplied the 
blood stream daily through the lymph-duct with a number of lymphocytes equal to 
more than half the total number in the body at any one time. If this is true, then the 
chief function of most of the white cells must be to increase the proteid content of the 
plasma. A similar question arises in cases with profuse pus formation, as cystitis, bron- 
chitis or bronchiectasis, since some of these patients are estimated to lose daily a number 
of white cells almost equal to the total number in their circulation at any one time. 

Fetal Blood. — In the 3-months' human embryo Engel found nucleated red cells of 
normal size and others larger, the " metrocytes of II Generation." These latter cells 
he describes as large spherical nucleated reds, 12 to 20ju in diameter, rich in protoplasm 
and with a relatively small nucleus 3.5 to 6/j, in diameter. (But some of these cells meas- 
ured from 17 to 20ju in size and had a nucleus measuring from 7 to 8/jl.) These numbered 
from 4 to 6 per 100 normal reds. (Metrocytes of I Generation he describes from mouse 
embryo's blood as spherical cells from 2 to 3 times the size of a normal red cell, the 
nucleus often in mitosis and rilling but a relatively small part of the cell; this, he says, 
is not a megaloblast nor a gigantoblast. At this stage there are no non-nucleated reds 

53 Zeitschr. f. klin. Med., 1904, Bd. 54, p. 232. 



508 CLINICAL DIAGNOSIS 

and no leucocytes.) At this stage are found 2 forms of normoblasts: those staining 
orange, from 5 to gfi in diameter and their nucleus 3.5 to 5^1; and those staining red 
(Ehrlich stain), about 7 to 8jjl in diameter, with a relatively large nucleus rich in structure 
5 or 6/j. large and their protoplasm scanty and ragged. Belonging to this latter group 
are some large cells i6/jl in diameter and a nucleusof 1 iji. These are Ehrlich'smegaloblasts. 

In embryos of 6 cm. length the non -nucleated reds were to the nucleated as 12 : 1 ; 
of 12 cm. embryo, 55 : 1; of 16 cm. 150 : 1; of 19 cm. 176 : 1. In the 6 cm. embryo the 
metrocytes were 4% of the reds; in the 12 cm., 0.25%, and latter none were found. The 
leucocytes in the 6 cm. embryo were to the reds as 1 : 500 to 1000. - 

Engel admits that embryos of the same age differ so that he could not tell the age 
of the embryo by studying its blood. 

We have had opportunity to study the blood (see Fig. 124) of a fetus 15 cm. long, 
and found: red cells 1,168,000; hemoglobin 25%; leucocytes 9000; nucleated reds 1 : 19 
of the total reds ; normoblasts and intermediates some showing beautiful polychroma- 
tophilia. 

In an embryo 20 cm. long we found: reds 2,652,000; leucocytes 28,000; hemoglobin 
38%. 

In an embryo of 23 cm. Engel found the reds (heart's blood) 3,300,000; hemoglobin 
80%; leucocytes 40,000. Nucleated reds were to non-nucleated cells as 1 : 120 and all 
red cells with nuclei were normoblasts. Of the leucocytes, the granular were to the non- 
granular as 2 : 5 ; neutrophile myelocytes and leucocytes and all transitional forms were 
present; also a few eosinophiles. 

The blood of a 27 cm. embryo contained nucleated and non-nucleated reds in relation 
of 1 : 200, leucocytes to erythrocytes as 1 : 90, polymorphonuclears to mononuclears 
as 4 : 5. 

LEUCOCYTOSIS 

By the term leucocytosis was formerly meant any increase of the white 
cells of the blood above the highest limit of normal, that is above 10,000 
per cmm. As the term is now used, however, the increase must be 
transitory and it must affect especially the polymorphonuclear finely 
granular cells. Ten thousand leucocytes per cubic milimeter may be con- 
sidered the upper limit of normal although some normal persons have 
for a long time a leucocyte count of from 10,000 to 12,000. Yet in judg- 
ing a leucocytosis we consider as limit more the count which would be 
expected in that particular person at that particular time ; that is, in the 
condition in which he then is. Many cachectic persons have for long 
periods of time very low counts e.g., 4000 cells, and a rise to 8000 would 
mean for them as much as a count of 20,000 would in a more normal per- 
son, for a leucocytosis represents a reaction, a struggle, and the result 
must be judged relative to the person making the struggle. One of our 
cases of typhoid fever had a leucocyte count of 1600. A parotitis de- 
veloped and the leucocytes promptly rose to 3200. This was a true 
leucocytosis for that person at that time. 

A leucocytosis also is transitory and symtomatic. This distinguishes 
it from leukemia. And, finally, the term leucocytosis is used if the rise is 
due to an increase of the polymorphonuclear neutrophile cells. If the 
increase involves the mononuclear non-granular cells, the term lympho- 
cytosis is used; if the polymorphonuclear eosinophiles, eosinophilic^, if the 



THE BLOOD 509 

mononuclear granular cells, mvelemia, etc. It is very seldom that i group 
of cells only is increased; usually other groups also are, but to a less degree. 
To some this would suggest that these groups are directly related, but a 
better explanation may be that several tissues respond to the same stimuli. 
Since the various cells may have little relationship to each other, that is, 
since i cell is not certainly a younger form of another, it is their absolute 
number which should be considered rather than their relative number, 
that is, rather than their percentages or " formula." The absolute number 
of a group of cells may increase even though its percentage diminishes 
providing the total count rises, while the reverse also is true that when the 
percentage seems to indicate an increase the absolute number may have 
dropped if there is a diminution of the total number. For this reason one 
should not make a differential count unless he makes a total count at the 
same time. 

That i group of cells may remain unchanged while other groups change 
much, is well illustrated by the case of Frazier and Halloway, in which the 
count was 13,040. Of these the polymorphonuclears were 78.2% (i.e., 
10,197) and the small mononuclears 16.8% (2101). The total count later 
rose to 54,960 of which the polymorphonuclears were 90.4% (49,684) and 
the small mononuclears only 4% (i.e., 2198 or the same as before). 

The general type of leucocytosis — i.e., one with an equal increase of all 
the types of leucocytes — is rarely seen. Such a leucocytosis does result, 
e.g., from stasis of blood in the capillaries, following a cold bath, or massage, 
while the digestive leucocytosis and that of pregnancy suggest this in some 
degree. 

Classification of the Leucocytoses (Limbeck). — 1. Physiological: 
(a) Digestion; (b) Pregnancy; (c) Newborn. 2. Pathological; (a) Inflam- 
matory; (b) Malignant tumors ; (c) Post-hemorrhagic ; (d) Agonal. 3. After 
medicinal and therapeutic measures. 4. Various other causes, as shock, etc. 

Physiological Digestion Leucocytosis. — If following a fast of 12 
or more hours a normal person partakes of a meal rich in proteid his leuco- 
cyte count usually will rise to about % above its normal number. The count 
should be made hourly. It begins to rise in about 1 hour, reaches a maxi- 
mum in from 3 to 5 hours and then decreases. While the increase involves 
the polymorphonuclear neutrophiles especially yet the small mononuclears 
are increased to some extent, in some cases considerably. For some persons 
a preliminary fast is not necessary; others do not show this leucocytosis 
at all (Limbeck thinks habitual constipation explains this failure). Chil- 
dren show it more markedly than adults and the well nourished more than 
the poorly nourished. It is greatest in the infant after his first meal of 
cow's milk. In the nursing infant it is said to be absent and hence the 
opinion (Moro) that when it does occur it represents a reaction against a 
foreign proteid. Since due to proteid diabetics show it well. It is important 
that the meal be unusually large since the leucocyte count may not change 



510 CLINICAL DIAGNOSIS 

and even may drop after a light meal. This leucocytosis cannot be demon- 
strated in herbivorous animals and with difficulty in man after a vege- 
table meal. 

This leucocytosis is supposed to be due to the positive chemotactic 
influence of the absorbed products of proteid digestion. Hofmeister sug- 
gests that a proliferation of the large masses of lymphoid tissue along the 
intestine, which accompanies the digestive processes, may be the cause, 
and in support of this calls attention to the coincident" increase in the 
mononuclear non-granular cells. This accompanying lymphocytosis is, 
however, not always present. 

Jaffe says that in children the leucocytosis is not dependent on the 
meal, but is periodic. 

The reverse relation is also true. Starving persons have a low leucocyte 
count. Succi, who fasted seven days, had a count of 86 1 per cubic milli- 
meter, while the insane with melancholia often have counts below 3000. 
On the other hand, well-nourished persons often have counts from 10,000 
to 12,000. From this it has been argued that the leucocytes play an im- 
portant part in the absorption,transportation and assimilation of food and 
so their number will depend much on the age and nutritional condition 
of the person. 

The attempt has been made to use the digestion leucocytosis as an aid 
in the differential diagnosis between pernicious anemia and cancer of the 
stomach. In severe blood diseases, as pernicious anemia, in ulcus ventriculi 
and in other gastric diseases a digestion leucocytosis can be demonstrated, 
while even in fairly early cases of cancer of the stomach it is sometimes, 
but not always, absent. It is absent in some benign gastric conditions. 54 
Gastric catarrh and involvement of the lymph-glands are given as the 
explanation for its absence. 

Leucocytosis of Pregnancy. — About 75% of women during the last 
months of pregnancy have a leucocyte count which is above normal, aver- 
aging about 13,000 per cubic millimeter. This is especially true of the primi- 
para and yet the explanation may be more her youth and better nutritional 
condition rather than the fact that she has had no previous pregnancies. 
The count rises until the end of pregnancy and then diminishes for from 
4 to 14 days after delivery. The differential count may remain practically 
normal and yet the polymorphonuclear neutrophiles are usually increased. 
In multiparas the leucocytes rise but usually within physiological limits. 

V. Limbeck considers the leucocytosis of pregnancy as a prolonged 
digestion leucocytosis, due to the need of additional nourishment for the 
mother and child. In favor of this is the observation that the count is not 
increased, is in fact even diminished, after a heavy meal. This is due, it 
is said, to a migration of leucocytes to the placenta where is the greatest 
accumulation of the positively chemotactic products of digestion. The 

54 Renchi, Arch. f. Verdauungskr., Bd. vii. 



THE BLOOD 511 

condition of the breasts is also suspected ; others ascribe it to an overactivity 
of the lymphatic system. But the view most commonly held now is that 
in pregnancy there is a slight intoxication against which a primapara 
reacts differently than a multipara. Thomson found that of 33 counts on 
12 pregnant women made during the 8 months of pregnancy but 1 was 
below 7000; the highest was 13,200. 

But the question is, What is the usual count for a normal woman? Is it 5500? If 
so, pregnancy causes in all cases a relative rise and in most an absolute leucocy tosis . 
Zangemeister and Wagner 55 think the question a complicated one. Of 47 normal non- 
pregnant women, from 21 to 34 years of age and all under practically the same condi- 
tions, 35 (74%) had counts above 10,000 (mean count about 12,500). The leucocyte 
counts of pregnant women (57 cases) varied within the same limits as the non-pregnant 
(70% above 10,000; mean count between 12,500 and 15,000), nor did the number of 
previous pregnancies seem to make any difference. During labor the count rose in nearly 
all of the 63 women examined even to 3 times the previous count, the maximum at or 
just after delivery. This was especially marked in cases of prolonged labor and of those 
who suffered greatly. In quick, easy labors the rise is insignificant. 

In 75 cases the count decreased during the puerperium rapidly to normal. On the 
seventh or eighth day there was an increase of mononuclears which accompanied the 
involution of the uterus. (Rouslacroix and Benoit). A study of 2 cases of version led 
them to think that the rise might be due to the contractions of the uterus. 

In pathological cases the leucocytes give no aid in diagnosis or prognosis, since the 
counts are no higher than some seen in the physiological capes. 

Lobenstein 56 believes that there is a genuine leucocytosis of pregnancy. He found 
that the average of 50 cases during the ninth month of pregnancy was 11,854 for primi- 
paras and 9346 for multiparas; and on the third day of the puerperium 13,200 for primi- 
paras and 11,600 for multiparae. These figures are too nearly normal to have great sig- 
nificance. The digestion leucocytosis test was tried in 20 cases and found present (p. 
510) in 13 while an actual diminution in the count was noted in 6. Of 13 cases of 
eclampsia, in 6 mild cases the highest count was 31,000; in 6 severe cases from 40,000 to 
50,000; and in 1 severe fatal case it was 106,000. He concludes that the leucocytosis 
runs roughly parallel to the degree of intoxication and to the resistance. A low count 
and a rapidly falling count are bad signs. 

Leucocytosis of the Newborn. — -Although the fetus has many 
blood-building organs yet the leucocyte count is very low since there is as 
yet no function for these cells (Askanazy) . The statement is often made 
that at birth the leucocyte count ranges from 17,000 to 21,000 and that 
after the first feeding it rises from 26,000 to 36,000, the increase involving 
the small mononuclears especially. On the first day after birth Gundobin, 
Carstanjen, and Warfield 57 found the average count about 26,000 
(11,700 to 34,700), while on the third day the average was 13,270 and 
on the eleventh day 15,740. For the first few days there is an absolute 
increase in the number of polymorphonuclear neutrophiles. Their per- 
centage was 70.42 on the first day, 53.16 on the third and 34.2 on the elev- 
enth. The large mononuclears and transitionals are high, being 10.76%, 

55 Deut. med. Wochenschr., July 31, 1902. 

56 Am. Jour. Med. Sci., 1904, vol. cxxviii, p. 281. 

57 Amer. Medicine, September 20, 1902. 



512 CLINICAL DIAGNOSIS 

16.67%, and 15.98%, respectively on these 3 days. The eosinophiles 
vary much ; Mastzellen and myelocytes are few. It is not until the eleventh 
day that the blood picture is that usually considered normal for infants, 
i. e. t with 40% small mononuclears, etc. 

This high leucocyte count has been explained as due to abnormal con- 
centration of the blood or to a digestion leucocytosis, but the more rational 
explanation is that it is the result of the rapid blood formation at that 
age. Although the blood of normal infants varies much, yet this rather 
high count may continue until the third or even the sixth year, after which 
time the blood picture of the adult prevails. During these early years the 
polymorphonuclear neutrophiles vary from 18 to 40% and the small 
mononuclears from 40 to 60% of the total number. Often there is a slight 
increase in the eosinophile cells. Such are the reports of several observers. 
These figures are not the invariable rule, however, as many a teacher dis- 
covers when he attempts to demonstrate a lymphocytosis to a class and 
uses a new-born baby's blood only to find the picture of a normal adult. 

Pathological Leucocytosis. — Leucocytosis of Inflammations and 
Various Febrile Diseases. — Most of the acute pyogenic infections and 
many of the acute febrile diseases are accompanied by a leucocytosis, due 
to an absolute increase of the polymorphonuclear neutrophile cells, which 
runs roughly parallel to the temperature and which depends for its existence 
and grade on the activity of the inflammatory process and on the condition 
of the patient. 

The following general statements may be made. Whatever its im- 
mediate cause, a leucocytosis represents a reaction of the individual to the 
disease. In those conditions which usually call forth a leucocytosis a high 
count means a vigorous reaction, little more, while a low count may 
mean a poor reaction, hence indicate a poor prognosis, but it also may mean 
that the infection is so mild that it can elicit little or no reaction. 

Diseases differ much in their ability to produce a leucocytosis; or, to 
put it in a different way, the body defends itself differently against certain 
diseases; by means of a leucocytosis in some and by a leucopenia in others. 
In acute lobar pneumonia, scarlet fever and the pyogenic infections there 
is usually a leucocytosis the degree of which runs roughly parallel to the 
intensity of the body's reaction, while the absence of a leucocytosis in mea- 
sles, malaria, and tuberculosis is of great importance in diagnosis. Some 
diseases may begin with a leucocytosis and end with a leucopenia. This is 
true of some cases of typhoid fever. Some develop a leucocytosis during 
the course of the disease, as typhus fever, some cases of influenza, and small- 
pox. Some diseases, as malaria, ordinarily without a leucocytosis may in 
severe cases develop one. But in all cases more depends on the organs 
infected by a given germ than on the germ itself. For instance, typical 
typhoid infection of the lymphatic apparatus of the bowel causes no leu- 
cocytosis but the invasion of the pleural cavity or of the periosteum by 



THE BLOOD 513 

Bacillus typhosis will do so promptly. Tuberculous adenitis causes no 
rise of the white cell-count but tuberculous pneumonia may cause a 
high count. 

In cases of local infection, as abscess formation, the leucocytosis is a 
symptom related to the fever and other toxic features and evidently is, 
like them, caused by, and its severity determined by, the amount of toxine 
absorbed; for, following an operation which allows free drainage, both 
quickly drop to normal. For much the same reason the leucocyte count 
runs quite parallel to the richness of the exudate in pus-cells. In general, 
the leucocyte count is no indication of the severity of the condition; a 
simple local felon may cause as high a leucocytosis as an appendix abscess 
and a fatal pneumonia as little as a small boil. 

It is not the formation of an inflammatory exudate alone which governs 
the leucocyte count, for some patients with free drainage of pus may daily 
lose enormous numbers of white cells (almost as many as are in the cir- 
culation at any one time) (see page 507) and yet show a normal count. 
This is well seen in some cases of chronic bronchitis, bronchiectasis, cystitis, 
in various bone and joint abscesses with discharging sinuses, in empyema 
after operation, etc. The agent causing the leucocytosis seems the same as 
that causing fever, for they usually run parallel. 

One would expect that a great loss of cells in an exudate would cause a 
diminution of the white cell-count in the blood and there are cases of spread- 
ing peritonitis in which this count does for a time fall. 

Among the conditions causing leucocytosis are: Acute lobar pneu- 
monia, the best studied example (page 640). 

Acute tuberculous pneumonia (page 636). 

Acute articular rheumatism (page 644). 

Diphtheria (page 634). 

Acute cerebrospinal meningitis caused a leucocytosis in all of 21 cases 
(Osier); in 4 it was over 40,000; the highest was 47,000. The leucocyte 
count is of no especial value in distinguishing the various forms of men- 
ingitis, since it is present also even in the tuberculous form. 

An acute follicular tonsillitis usually causes a leucocytosis. This 
was true of 18 of 26 of our recent cases. (In 12 the count ranged from 
10,000 to 15,000; in 3 it was above 20,000; the highest was 27,000.) The 
temperature was high in all the cases with high counts. 

Scarlet fever (page 634). 

Mumps. The occurrence of a leucocytosis is disputed. 

In whooping- cough the leucocytes, especially the lymphocytes, are much 
increased, the, counts averaging 40,000. This leucocytosis is more pro- 
nounced the younger the child is. Its early appearance makes it of 
great value in diagnosis. It begins during the catarrhal stage and, con- 
tinuing through the paroxysmal stage, reaches its maximum during con- 
33 



514 CLINICAL DIAGNOSIS 

valescence. The increase of the count would seem to be due to an increase 
of the lymphocytes especially but others claim it to be a true leucocytosis. 

In Norton and Kohman's case of anthrax the leucocyte count was 
31,000 per c.mm. of which 81% were polymorphonuclear neutrophiles, 17% 
small mononuclears and 2% large mononuclears. 

Rabies sometimes causes a true leucocytosis of even 25,000, with 98% 
of the cells polymorphonuclear neutrophiles. 

Erysipelas causes a leucocytosis which runs fairly parallel to the tem- 
perature. The count ranges between 10,000 and 20,000 in mild and be- 
tween 20,000 and 30,000 in more severe cases. Its polymorphonuclear 
neutrophile character is more marked in adults than in children. These 
cells may reach 92% of the entire count in fatal cases. As the count falls 
the eosinophiles may rise considerably. 

In 6 of our cases the leucocytes were normal in 2, moderately elevated in 2 and were 
26,000 and 34,500 in the other 2. The red cells were normal in all. 

In acute ulcerative endocarditis the leucocyte counts are high as a rule, 
especially in those cases which run a protracted course. But a leucocytosis 
is not constant. In some mild cases and in rapidly fatal cases there may 
be none. 

In 6 recent cases at death the counts were 7070, 12,300, 13,600 (it had fallen from 
34,000), 17,000, 47,000 and 48,000 (it had risen from 9800). 

Acute Poliomyelitis. — In acute poliomyelitis Peabody, Draper and 
Dochez 58 found a constant and marked leucocytosis. They also found a 
constant increase in polymorphonuclear cells of from 10 to 15% and a 
diminution of lymphocytes of from 1 5 to 20%. 

In intestinal obstruction the leucocytes rise rapidly, to about 16,000 when 
the obstruction is partial, and to 20,000 or more when it is complete. If 
the count reaches over 20,000 cells during the first 24 hours the chances are 
that gangrene of the bowel has developed. This rise of leucocytes may be 
of value in a case of suspected post-operative obstruction (Bloodgood) . 

The myxedema which follows a thyroidectomy may be accompanied by 
a leucocyte count of even 49,000. 

Smallpox (page 634). 

Cholera. — During the algid stage of cholera the leucocytes may number 
from 40,000 to 60,000. The count rapidly falls during the stage of reaction. 

Pyogenic inflammations, not due to Bacillus tuberculosis, of the serous 
membranes, the meninges, pleura, pericardium and peritoneum, are accom- 
panied by a leucocytosis which bears some relation to the cellular richness 
of the exudate but more to the fever. The count varies with the prcgress of 
the disease since it may drop to normal while the process is stationary even 

58 Monograph of The Rockefeller Institute for Med. Research, No. 4, New York, 
1912, 97. 



THE BLOOD 515 

though the temperature remains elevated and then a slight spreading of the 
process will cause a rapidly rising count. This is well seen in pelvic in- 
flammations. 

In 99 cases of PLEURISY WITH EFFUSION the red cells were practically normal; 
in 65 of these the leucocyte counts were below 10,000 cells and in but 3 of the remaining 
34 cases were they over 15,000. Cabot reports almost exactly the same figures for the 
Massachusetts General Hospital (314 cases; 33% above 10,000; 6% above 15,000). The 
low counts are interesting since so many such cases are clearly tuberculous. 

In a series of cases all of which were possibly tuberculous the counts were found 
above 12,000 in 18.9%. In those of this series which were positively tuberculous it was 
above 12,000 in 9%; in those cases of combined pulmonary tuberculosis and pleurisy 
with effusion it was above 12,000 in 21.8% while, finally, in those cases of pleurisy with 
effusion combined with pneumonia it was above 12,000 in 78.5% of the cases. 

Empyema. — An inflammatory leucocytosis is the rule in cases of primary 
pneumococcus empyema and of those cases which follow acute lobar 
pneumonia. In such cases the leucocytosis is due to an increase of the 
polymorphonuclear finely granular cells and the white cell count curve 
will run roughly parallel to that of the temperature. In cases of pneumo- 
coccus empyema following pneumonia it is the continuous elevation of the 
leucocyte count together with the slight fever which leads to the correct 
diagnosis. This leucocytosis is valuable in excluding serous effusions. In 
the empyema prevalent during the influenza epidemic of 191 7-19, however, 
a very different state of affairs existed since the count usually was that of 
the preceding pneumonia even though this was below 3000 cells. 

In pneumococcus cases allowed to remain without operation for a 
long time the count may return to normal. 

Fibrinous Pleurisy. — In fibrinous pleurisy a slight secondary anemia is 
to be expected. The leucocytes in 37 cases which we reported ranged, in 
24, from 10,000 to 22,900, while in 13 the counts were normal. Lord found 
the count above 12,000 in 30% of his cases of fibrinous pleurisy. Since the 
count was above 12,000 in but 9% of his cases of pleurisy with effusion he 
suggests that the higher count in fibrinous pleurisy is evidence that this 
condition is more often a pyogenic infection than is the other. 

Influenza is a term which has been applied to a variety of conditions of 
chest, abdomen or nervous system due to organisms of the streptococcus and 
pneumococcus groups, to Bacillus influenzas, etc., which have had this in 
common, however, either that the condition was epidemic or if sporadic that 
the infection was particularly prostrating. The epidemic of 1917 to 1919 
proved quite satisfactorily that the organism of the disease is as yet un- 
known, that its infection produces a marked leucopenia and that it prepares 
the body for secondary invasions by the above-mentioned organisms which 
explain the various complications which give the more prominent character- 
istics to the disease, including often a leucocytosis. 

If we accept the diagnoses of the past, the leucocytes are normal in 



516 CLINICAL DIAGNOSIS 

about % of the cases (Cabot) and moderately increased in the rest. Some 
have stated that in the typhoidal or abdominal form of influenza there is 
leucopenia. 

In almost half of our sporadic cases the count was abcve 10,000, even 
25,000, at the height of the disease. Nearly all the cases in which several 
counts were made showed early a very low count, from 3,000 to 5,000, even 
when the temperature was from 100 to 105 , doubtless the count of the 
influenza itself, then a sharp rise due to a secondary infection. 

During the epidemic of influenza of 19 18-19 a leucopenia was the rule, 
the counts averaging about 4,000 cells, while in some cases even in the 
presence of extensive pneumonia the count was as low as 2,800. In other 
cases a leucocytosis was a feature of the pneumonia and in 1 case of 
empyema the count reached 80,000. 

The pyogenic processes of mucous membranes which cause fever are usu- 
ally accompanied by a leucocytosis, as enteritis, urethritis, etc. 

In acute bronchitis, the leucocytosis continues as long as the fever. The 
counts were between 10,000 and 20,000 in 30 of our 67 cases. 

In chronic bronchitis the emphysema and attending cyanosis may 
explain the occasional leucocytosis which was present in just half of our 
cases. The red cell counts averaged high, the mean being 5,000,000. Of 
25 cases in 3 the counts were above 7,000,000 (maximum 7,900,000). 

In a case of true foetid bronchitis, the leucocyte count was 22,500. 

In 11 cases of bronchiectasis the leucocyte counts were 20,000 in 2; be- 
tween 10,000 and 20,000 in 4; and normal in 5 afebrile cases. 

Among the local pus processes in which the leucocyte count is of value 
in diagnosis are appendicitis (page 644), pelvic inflammatory disease, 
abscess of the liver, emphysema of the gall-bladder, ovarian abscess, abscess 
of the brain, etc. 

In abscess of the lung counts as high as 60,000 have been reported. In 
our 3 cases they were 8,100, 12,300 and 12,500. In 2 cases of gangrene of 
the lung they numbered 20,000 and 48,000. 

In 25 cases of gonorrheal arthritis the mean count of red cells was 
4,500,000; the lowest was 3,600,000. The mean leucocyte count was 9 ,000. 
In 8 of 23 cases the white counts varied between 10,000 and 20,000. 

In perirenal abscess, 5 cases, the leucocyte counts were between 19,000 
and 26,000; pyelitis, 4 cases, between 10,600 and 19,500; pyelonephrosis, 
2 cases, 18,000 and 28,500; hydronephrosis, 2 cases, 6,400 and 9,000; 
and pyelonephritis, 1 case, 8,000. In renal calculus, 4 cases, the leucocyte 
counts during the colic were between 12,000 and 18,000. 

In gout the red cell counts were 5,000,000 or over in all but 2 cases 
(of 13 cases the lowest was 4,300,000). The leucocyte count rises at the 
onset of an acute joint attack. (In 18 cases there was a mild leucocytosis 
from 10,000 to 14,000 in 7 cases.) The variations in the leucocyte count 
run parallel to the temperature curve and the joint symptoms. 



THE BLOOD 517 

There is a leucocytosis in diabetic coma and in uremia. 

In dementia prcecox 59 there is a polymorphonuclear neutrophile leuko- 
cytosis of 15,000 or less coincident with the onset cf the abnormal mental 
phases for which no satisfactory explanation has been given. In general 
paresis there is often an absolute lymphocytosis, the total number of leuco- 
cytes ranging between 7,000 and 10,000, from 35 to 55% of which are small 
mononuclears. 60 

Following operations there is often a slight post-operative leucocytosis, from 
10,000 to 20,000, which is not due to infection, 6l which reaches its maximum 
in the first 12 hours after operation, and which is not accompanied by par- 
allel changes in the temperature or the pulse rate. This leucocytosis con- 
tinues for not over 36 hours. If, however, the count rises more than 10,000 
above what it was before operation and remains elevated for over 2 days 
one should suspect a post-operative pyogenic complication. The highest 
count in our series which followed a nephrotomy was 32,000 cells. While 
the nature of the operation seemed to have little influence on the count, 
yet the rise did seem to run roughly parallel to the amount of tissue trau- 
matism produced. There is little relation between a post-operative 
leucocytosis and a post-operative fever due to infection. Chloroform 
anesthesia, but not ether, can cause a true but transitory leucocytosis. 

No sharp line can be drawn between the leucocytosis of infected and of 
non-infected wound repair except that the latter is on the wane at a time 
when the former is just beginning. 

When the packing of a wound is changed the leucocyte count may rise 
somewhat. In the case of a closed wound the leucocyte count is a good 
index of an infection. 

The diseases which cause as a rule no leucocytosis are influenza (page 
516), typhoid fever (page 639), measles (page 633), and tuberculosis 
(page 634). 

Pseudoleucocytosis. — Certain blood-changes other than a rise of the 
white count sometimes occur under conditions which usually cause a 
leucocytosis and are supposed to have much the same significance. Among 
these are iodophilia (page 524), degenerations of the leucocytes, fragmen- 
tation of their nuclei (as occurs in cancer), the appearance of myelocytes 
and a relative increase of the polymorphonuclear neutrophiles although the 
total count does not rise above normal. This is met with in cancer, septi- 
cemia, etc. 

Leucocytosis of Malignant Disease. — Cases of carcinoma (page 650) 
and of sarcoma especially (page 652) are frequently accompanied by a 
leucocytosis of polymorphonuclear cells which disappears after the removal 

59 Barnes, Am. Jour. Insan., April, 1909, vol. Ixv. 

60 Cornell, Am. Jour. Insan., July, 1907, vol. lxiv. 

61 Frazier and Halloway, Contrib. from the Wm. Pepper Lab. of Clin. Med., 1902 
No. 3. Am. Jour. Med. Sci., September, 1902, vol. cxxiv. 



518 CLINICAL DIAGNOSIS 

of the tumor. This is frequently true of cancers of the internal organs, 
especially of the stomach, but not of epithelioma of the skin. This leuco- 
cytosis bears little relation to the situation of the tumor unless it metas- 
tasizes to the bone-marrow, in which case it may simulate leukemia. 

Post-Hemorrhagic Leucocytosis. — Large hemorrhages are often fol- 
lowed by a leucocytosis which begins in from 10 to 15 minutes and which 
in 1 hour may reach from 16,000 to 18,000. It lasts a few days and then 
disappears. The increase is of the polymorphonuclear neutrophile cells. 
Its cause cannot be a new production of cells, since it begins too suddenly. 
Some attribute this to the tissue lymph which flows into the vessels in order 
to restore the volume of blood, carrying with it a large number of white 
cells (but these cells should be small mononuclears) , but the nature of the 
wound is more important, since an injury without hemorrhage can cause a 
leucocytosis and even a severe hemorrhage resulting from a very slight 
injury (e. g., that from a gastric ulcer) a slight leucocytosis lasting in some 
instances but 2 days. Stassano and Billou 62 found that a leucopenia may 
follow severe hemorrhages and a true leucocytosis smaller ones. 

In a case of cirrhosis of the liver with fatal hemorrhage from the 
stomach the red cell count before death was 1,960,000, hemoglobin 23% and 
the leucocytes 23,000. 

Agonal Leucocytosis. — Since belief in an agonal leucocytosis antedates 
our knowledge of inflammatory leucocytoses it is more than likely that the 
leucocytoses of pneumonias were included under this heading. And yet 
this would not explain all of the high ante-mortem leucocyte counts, e. g., 
Cabot's case of pernicious anemia which resembled a leukemia. Such cases 
are rare, it is true. In most diseases the leucocyte count does not 
change or even drops at death, while the rises which one does find are 
ascribed by Ehrlich to the slowing of the circulation and hence to the ac- 
cumulation of the leucocytes along the periphery of the blood-vessels. 

Medicinal Leucocytosis. — The administration by mouth or subcutane- 
ously of any one of a very long list of drugs, including the ethereal oils, 
tonics, myrrh, turpentine, peppermint, etc., may cause a definite leuco- 
cytosis. If the drug is administered by mouth the result would seem com- 
parable to a digestion leucocytosis, while if injected subcutaneously the 
local reaction also may be important. It is of interest that the extracts 
of certain body tissues also seem positively chemotactic to leucocytes. 

The reverse also is true, for blood poisons which destroy the cells, as 
phenacetin, the chlorates and pyrogallic acid, may cause a drop in the 
leucocyte count. 

Other Causes of a Leucocytosis. — In the case of animals simple violence 
will cause a leucocytosis. In man, hard work, severe sweating, heat and 
cold, many vasomotor influences, anything which slows the circulation and 

62 Compt.-rend. Soc. Biol., 55, p. 180. 



THE BLOOD 519 

causes cyanosis will do the same. Thayer found that in typhoid fever 
cold baths, especially these which made the patient shiver, would cause a 
rise of about 6000 cells; the formula remained the same. Violent exercise 
will cause a leucocytosis ; for example, the leucocyte counts of the runners 
of a 2 5 -mile race rose from 14,000 to 22,000. 63 

Mixed Leucocytosis. — By the term mixed leucocytosis was formerly 
meant an increase of the ameboid and non-ameboid cells ; that is, of the gran- 
ular and non-granular cells. But the discovery that the latter are ameboid 
robbed this term of all significance. Others used it of a leucocytosis with 
the presence of neutrophile myelocytes. The best illustration of this is 
myeloid leukemia in which condition the absolute number of myelocytes 
may vary from 50,000 to 100,000 per cubic millimeter. In no other con- 
dition does the absolute number of myelocytes in the blood rise above 1000 
(Ehrlich) . In leukemia appear the largest number of eosinophile myelocytes 
seen, of Mastzellen also, together with all forms of the non-granular cells. 
The next most important condition in which a mixed leucocytosis may arise 
is pernicious anemia. In focal infections or malignant nodules of the bone- 
marrow the blood may appear even leukemic. 

And yet almost any leucocytosis could be called "mixed" since the 
higher the count the younger are the forms which appear in the blood. 
Especially is this true of children whose blood is very unstable, and particu- 
larly of those with diphtheria, anemia, rickets, congenital lues and pneu- 
monia. In the latter condition especially, one may find just after the crisis 
myelocytes and megaloblasts. Myelocytes have no significance in the 
blood of adults if they disappear as the count falls, but should they still 
appear after the count falls their presence might mean exhaustion of the 
bone-marrow. A mixed leucocytosis is not rare in severe post-hemorrhagic 
anemias. 

Mastzell Leucocytosis. — The only condition in which the Mastzellen 
of the blood are always increased is splenomyelogenous leukemia. In that 
disease even 1 5 or 20% of the leucocytes may be these. Isolated cases with 
an increase of these cells in the blood have been reported, as of cancer, 
septic bone disease, various skin diseases and even chlorosis. The diffi- 
culty of recognizing these cells in specimens stained with methylene blue 
mixtures should be emphasized. 

Leucocytosis with an Increase in Endothelial Leucocytes. — The endo- 
thelial leucocytes are increased absolutely in typhoid fever, measles, and 
especially in malaria in which disease even 20 to 30% of an almost normal 
count may be these cells, a point of great value in diagnosis. 

Lymphocytosis. — Ehrlich classified leucocytoses as active, passive and 
mixed. The ordinary leucocytosis is "active " because it is the actively 
ameboid cells which are increased. These are supposed to have entered the 

63 Larrabee, Jour, of Med. Research, 1902, vol. ii. 



520 CLINICAL DIAGNOSIS 

circulation in abnormal numbers in response to a positive chemotactic agent. 
Ehrlich called a lymphocytosis " passive " because he supposed these cells 
to be passively and mechanically washed out of the lymphatic tissue by 
the circulation. 

The lymphocytes are ameboid on a rather hot stage (44 to 46°C). 
They migrate into the tissues in certain skin diseases, also in tuberculosis, 
lues and malignant disease. They are the cells of the tuberculous pleural 
exudates. Under these conditions they certainly are ameboid. 

Rous 64 found that in man a lymphocytosis may follow active exercise 
unless this is very severe (as a 2 5 -mile run) in which case there may be a 
decrease in the absolute number of mononuclear cells of the blood. This 
increase is not due to an increased flow of lymph since the increase of these 
cells in the blood is even twice that in the thoracic duct and is a greater 
addition than any simple lymphagogue, as glucose, can produce. 

These cells contain a lipase capable of splitting wax and fat to glycerine 
and fatty acid, which is interesting since the bodies of tubercle bacilli 
contain about 30% of waxy substances. 

Rous found that normally the lymph of the thoracic duct furnishes the 
blood with a large proportion of its lymphocytes and that under normal 
conditions this supply is quite constant. 

Physiologically, a lymphocytosis exists in infants and during a digestive 
leucocytosis. The mononuclear cells are definitely increased at high alti- 
tudes, averaging 43.6% (polymorphonuclear neutrophiles 54%). 65 Patho- 
logically it occurs in the simple gastro-intestinal disturbances of children 
(page 647), in whooping-cough (page 513), in cervical adenitis, during the 
reaction to tuberculin, in malignant lymphomata and in sarcoma multiplex 
cutis. The greatest increase is in lymphatic leukemia in which disease over 
90% of the 140,000 or more cells may be mononuclears. They are abso- 
lutely increased in splenomyelogenous leukemia, while after a splenectomy 
they may slowly increase to even twice their normal number, which increase 
begins about a month after the operation and continues for even 1 year. 

The best illustrations of the chronic diseases with a lymphocytosis are 
hereditary lues and severe rickets. The statement is often made that a 
lymphocytosis is common in chlorosis, pernicious anemia, debility, late 
typhoid fever, Graves' disease, hemophilia, scurvy and during thyroid 
treatment, but as a rule the increase is only relative. 

The leucocytosis of children is often chiefly a lymphocytosis, e. g., 
the case with enlarged tonsils mentioned by Churchill, with a count of 
20,000, 70% of which were small mononuclears. In the diagnosis of lym- 
phatic leukemia these cases should be remembered. The total leucocyte 
count may be low and yet a true lymphocytosis exist, as in a recent case of 

64 Jour, of Exp. Med. 1908, vol. x; Proc. of the Soc. for Exp. Biol, and Med. 1907, 
vol. iv. 

65 Webb and Williams, Trans, of the Fifth Annual Meeting of the National Assoc, 
for the Study and Prevention of Tuberc., p. 231. 



THE BLOOD 521 

amebic dysentery with a count of 2,500 cells 68% of which were small 
mononuclears. 

Leucopenia, or a count below 5000 cells per c.mm., may result from the 
reduction in 1 group of cells or from a general reduction of all groups. The 
former is seen during typhoid fever. A leucopenia is said to be the first 
stage of a leucocytosis. In tuberculosis of lymph-glands the count may be 
even below 600 cells. In aplastic anemias it may fall as low. 

Ehrlich mentioned an interesting case of general lymphosarcoma with 
but 0.6% of small mononuclears in the blood. 

Conditions with leucopenia have been reported under the name "alym- 
phaemic lymphomatosis." Such a case was reported by Schwartz with 
acutely swollen glands and fever whose leucocyte count was only 600 per 
c.mm. and all of them lymphocytes (no autopsy) . 

The relationship between leucopenia and infection of the abdominal 
organs deserves mention. Infections with Bacillus typhosus usually pro- 
duce a leucopenia when the local infection is chiefly abdominal, but they pro- 
duce a leucocytosis when other organs as pleura, lung, etc., are involved ; 
tuberculous peritonitis and " abdominal influenza " cause no leucocytosis. 

The leucocyte count may fall to a low figure during the convalescence 
of some fevers. At the end of typhoid it is often as low as 2000 cells. 

In a recent case of hemoglobinuria the red cell count was 2,500,000 and 
the leucocyte 950 per c.mm. 

Cases of starvation or malnutrition from any cause have a low leucocyte 
count; for example, voluntary starvation (page 597) and that due to disease 
as cancer of the esophagus (page 651). In one of our cases of ulcerative 
colitis the patient's red count was 2,100,000, and the leucocytes 700 per 
c.mm. In influenza even with pneumonia the count may drop to 2800 or 
lower. In measles there is often leucopenia. 

In 1 of our cases of measles, No. 9621, a man 22 years old, the count 
was 3600 per c.mm. at the height of the disease. 

In acute miliary tuberculosis (page 637) the counts are often very low. 
In all chronic intoxications as alcohol, morphia, lead, ether, mercury, ar- 
senic (hence the drop in leukemia?), leucopenia is the rule. Benzol will 
produce a marked leucopenia. 

Eosinophilia. — By eosinophilia is meant an absolute increase of the 
eosinophile cells in the circulating blood. The normal percentage of these 
cells varies from 2 to 4% and many use this term of conditions in which the 
percentage is increased; but the term should be limited to those cases in 
which the absolute number of these cells is above 250 per c.mm. 

Those conditions in which these cells are increased are so varied that 
they have been well termed the most capricious cells of the blood. 

I. There is a physiological eosinophilia during childhood. 

II. Diseases of the Hematopoietic Organs, i. Bone-marrow — 
Diseases of the bone-marrow often produce an eosinophilia. (a) In spleno- 



522 CLINICAL DIAGNOSIS 

myelogenous leukemia there is usually but not always an absolute increase 
of these cells, even 29,000 per c.mm. (Zappert). In a few cases they have 
been reported absent. In (b) sarcoma of the bone-marrow, page 652, 
(c) osteomyelitis and (d) osteomalacia they are sometimes present in 
great numbers. 

2. Spleen. — About 1 year after the removal of the spleen there slowly 
develops an eosinophilia which lasts for several months. These cells are 
increased to from 30 to 50 times their normal number and may make up 
even 36.6% of the total leucocyte count. A somewhat similar condition 
is present in cases with chronically enlarged spleen, in which the percentage 
of these leucocytes may vary from 7 to 12, and in new growths of the spleen. 
The reason may be that these spleens are functionless. 

3. Lymph- glands . — That an eosinophilia may result from disease of the 
lymph-glands is not certain. In support of such conditions have been cited 
cases with extensive metastases of carcinoma to these glands and eosinophilia 
but metastases to bones also were not excluded. In one case with such bone 
metastases the eosinophile cells numbered 60,000. 

III. Asthma. — During the paroxysms of true bronchial asthma from 
10 to 20% and in a case of Billings between 53 and 54% of the leucocytes 
may be eosinophile cells. This has considerable value in the differential 
diagnosis between true asthma and asthmatic attacks due to other causes. 
In emphysema these cells are also increased, in 1 case numbering 53.6% of 
a total of 8300 leucocytes. 

IV. Skin Diseases. — Many skin diseases produce an eosinophilia, which 
seems to depend more upon the extent of the lesion than on its nature. 
The cells in the pustules of some conditions are sometimes all eosinophiles. 

In a case of pemphigus reported by Zappert there were 4800 eosinophiles 
per c.mm. of blood; in 1 of pemphigus vegetans of the Baltimore clinic 
the total count was 20,400 leucocytes, 2.6% of which were eosinophiles, and 
on another day 11.6%; in pellagra and psoriasis these cells are sometimes 
increased; in urticaria even 60% of the total count may be these cells; 
in a case of the Baltimore clinic of purpura with cyanosis 11% of 52,000 
leucocytes and later 25% of a count almost as high were eosinophiles; 
in certain cases of eczema they are increased; of 2 cases of scleroderma in 
this clinic, in 1 they constituted 2.4% of 7000 and in the other 3.3% of 
10,500 leucocytes; in 5 cases of purpura simplex the red cells and hemo- 
globin were practically normal but 3 had a leucocytosis of from 10,100 to 
40,600 of which from n.3to25.6% were eosinophiles. (In 1 of these three 
cases, one with myositis the eosinophiles were 12.1% of 40,600 cells; in 
another 25.6% of 20,3 00.) Iniof3 cases of purpura rheumatica there was a 
leucocytosis of 13,300; in 1 of 3 of Henoch's purpura they were 10,000. 
Of 2 cases of purpura hemorrhagica in 1 the eosinophiles numbered 4-5% 
of 6600, in another 9.4% of 5200. In 1 case of morbus erronum 18% of the 
5000 leucocytes were eosinophiles. Later they were normal. Chemical 






THE BLOOD 523 

irritation of the skin, for instance by mercuric chloride, may increase these 
cells even to 14%. 

V. Parasites, — Any animal parasite, from the harmless pin-worm to 
the most malignant uncinaria, may cause an eosinophilia, but it does not 
always, nor does its degree bear any relation to the severity of the infection. 

In amebic dysentery of children especially there may be a slight eosino- 
philia. 

In 1 of our cases of amebic dysentery, No. 8174, a man 36 years old, these cells were 
8% of a total count of 8,800. 

Brown demonstrated this as a most important point in the diagnosis of 
trickiniasis. In 1 of his cases 68.2% of the 35,000 leucocytes were eosino- 
phils . This eosinophilia is not always present in trichinosis as in one case 
reported by Howard and another by Da Costa, but these cases are rare. In 
Brown's case they fell gradually to normal. The neutrophile cells in these 
cases are relatively and absolutely low. Brown was unable to obtain any 
Charcot-Leyden crystals from the blood and so suggests that some other 
element than an increase of eosinophile cells is necessary for their presence. 

In uncinariasis the percentage of eosinophile cells usually ranges from 
8.2 to 10 and in 1 case reached 72% of the count. Our highest count was 
13% of a total of 7400 leucocytes. In 1 case infected with Taenia saginata 
the eosinophile cells were 34% of the total count; of Ascaris lumbricoides, 
19%; Oxyuris, 16%; Strongyloides intestinalis, 13.5%; Bilharzia, 20%. In 
filariasis they vary from 4 to 17% but in Calvert's case they reached 22% 
during the day (in others the maximum is at night) . Calvert thinks that 
the grade of the eosinophilia depends on the acuteness of the attack and 
so it may be absent in long-standing cases. He found that the number of 
these cells in the circulation bears an inverse relation to the number of 
embryos, the former increasing during the day when the embryos disap- 
pear. In the diagnosis of hydatid cysts of the liver an eosinophilia is con- 
sidered important. It varies from 7 to 20% and in 1 case reached 40%. 
During the afebrile stage of malaria these cells have risen to 20.4%. 
In dracontiasis they are reported as from 6.4 to 36.6%. 

VI. A post-febrile eosinophilia may develop after many fevers but in 
most cases the increase is only to the upper limits of normal. In scarlet 
fever these cells may vary from 8 to 15% during the course of the fever but 
in all other fevers they first diminish and then rise as the temperature drops. 
After the crisis in pneumonia they may number 430 (5.7%) ; in acute arti- 
cular rheumatism, 970(9.4%) ; in malaria 1 day after the chill, 1486(20.34%) ; 
in varicella they may rise to 16% ; in measles to 5% ; and in rickets to 20%. 

VII. During a positive tuberculin reaction these cells may fall and then 
rise even to 3220 (26.9%). In 1 case reported by Grawitz their absolute 
number was 41,000 of a total of 45,000 leucocytes. 

VIII. These cells are often increased in diseases of the genital organs; 



524 CLINICAL DIAGNOSIS 

in all ovarian diseases except cancer, in 10 of 18 cases with ovarian cysts and 
abscesses and in gonorrhea, especially posterior urethritis and prostatitis. 

IX. In malignant disease the eosinophiles sometimes make up from 
7 to 10% of the leucocytes while in one case of lymphosarcoma they num- 
bered 60,000 cells. 

X. After certain medicines, as camphor (in which case they may rise 
to 9%) or the inhalation of carbon dioxide, the eosinophiles are sometimes, 
though rarely, increased. 

XL In diseases of the sympathetic nervous system. 

In the diagnosis of animal parasites, of asthma and of diseases of the 
bone-marrow the count of the eosinophile cells may be of great value. 
These cells certainly appear in the blood and tissues in response to some 
specific positive chemotactic agent. The best recent review of this subject 
is that of Howard. 66 We find in some pus, some sputa, etc., practically 
only eosinophiles, although the differential count of the circulating blood 
of these patients may be normal. 

It is important that in the blood of some rare cases we find a group of 
leucocytes which might be either neutrophiles or eosinophiles (see page 499). 
Were this the testimony of but few their technic or their judgment might 
be suspected, but several have reported such cases. 

Iodophilia. — By iodophilia is meant the presence in the blood of 
leucocytes whose protoplasm either takes a brownish red color or contains 
granules which take that color when the specimen is treated with iodine 
(the ' 'intracellular reaction") or the presence in the plasma of similarly 
stained granules from 2 to 8/z in size (the " extracellular reaction"). 

If normal blood is thus treated all the blood elements will take a bright 
yellow stain. 

The reagent used contains: iodine, 1 gm.; potassium iodide, 3 gms.; 
water, 100 cc. ; and gum arabic, 5 gms. A drop of this solution is placed on 
a slide and an unfixed smear of fresh blood is pressed down into the drop. 
The excess of stain is removed with a piece of filter paper applied at the 
edge of the cover-glass while this is held firmly against the slide. In judging 
the degree of iodophilia both the number of the cells and the degree to which 
these are affected are considered. Since all leucocytes contain some glycogen 
one must assume in iodophilia a special degeneration of those leucocytes 
which give the reaction, perhaps of toxic origin, which increases their 
affinity for iodine. 

This reaction is positive in a great variety of conditions. Locke con- 
siders the test independent of but of nearly equal value with a leucocytosis. 
It is met with in all the anemias except chlorosis, in leukemia, in nearly all 
cases of septicemia, especially those with leucocytosis, in cases with purulent 
exudates and especially in pneumonia. It is invariably present in severe 

66 The Jour, of Med. Research, Dec., 1907. 



PLATE II 

A, B, and C are groups of cells from three cases of Acute Lymphatic Leukemia. 

A. Cells from a case of the large-celled variety. 

B. Cells from a case of the small-celled variety. 

C. In this case the cells were almost achromatophilic, with the protoplasm slightly acidophilic. 

Leucocytes from Normal Blood and Malaria (in which condition is a large number of large 
mononuclear nongranular forms). 
9, io, 13, 19. Large mononuclear cells. 

11. A giant mononuclear cell. 

12. A mononuclear cell with the Wolff-Pappenheim granules. 

14. An eosinophile leucocyte. 

15. A naked nucleus. 

16, 17. Small mononuclear cells. 
18. Mastzell. 

20. A neutrophile leucocyte. 

21. Trypanosoma gambiense. 

22, 24, 25. Red cells with Grawitz's basophile granulation. 
23. A blood platelet. 



PLATE [] 







LYMPHATIC LEUK/EMIA. 












18 




^ 



LEUCOCYTES OF NORMAL BLOOD AND MALARIA. 









TRYPANOSOMA GAMBIENSE, 
FROM A CASE OF "SLEEPING SICKNESS. 
STAINED WITH HASTING'S MODIFICATION OF 
ROMANOWSKI'S STAIN. ALL DRAWN TO SAME SCALE. 






BASOPHILE GRANULES 

OF RED BLOOD CELLS. 

F. S. Lockteood. 






THE BLOOD 525 

septic conditions 67 and is valuable in this diagnosis if there is no leuco- 
cytosis. Another claim made is that this test is negative in cases of ovarian 
cyst with twisted pedicle and in other conditions without pus formation, 
even though there is a high leucocytosis. 

BLOOD PLATELETS 

Blood Platelets, Blutplatchen, Plaques (Kemp, Osier), Hematoblasts 
(Hayem), (Plate II, 23). In the blood are the so-called ''third corpuscles," 
small bluish bodies without nucleus or cell membrane and containing no 
hemoglobin, about 3/z in diameter, round, oval, or rod-shaped, according to 
the view-point and not biconcave. In an ordinary fresh blood preparation 
they have a peculiar bluish refractivity like the protoplasm of a non- 
granular leucocyte, but they stain like nuclear material. Platelets when 
perfectly fresh are slightly granular, but when removed from the blood- 
vessel they at once become hyaline, glassy and very sticky, then pale 
and disappear or they unite in masses of 2 to hundreds and disintegrate 
rapidly, even in a few seconds forming the so-called Schultze's ''granular 
masses" (see Fig. 125), from the periphery of which radiate fibrin strands 
and at the edges of which are vacuole-like areas, the so-called ''viscous 
metamorphosis" of Eberth or the 'mucoid degeneration" of Osier. 

In normal blood specimens stained with the usual Romano wski 
mixtures, they are seen in groups of from 1 to 10. They would seem to 
consist of protoplasm, sometimes hardly visible and sometimes definitely 
stained and swollen almost to the size of a red corpuscle 68 , and of nuclear 
material in rows of blue or reddish dots sometimes arranged in a spherical 
mass. 

When the platelets rest on cells they may closely resemble malarial 
parasites. 

Their size in the fresh specimens varies from 2.5 to $/j, in diameter 
(Determann); 1.5 to 3.2,11 (Osier); 2 to 7m (Preisich and Heim). In general 
their size varies inversely as their number; that is, the more the platelets 
the smaller they are. Their fragility also is more marked when they are 
increased. It is best to study them at a temperature not over 4o°F., for 
then their changes are much slower, requiring minutes instead of seconds. 
Some soon show clear areas, either in the center or on one side, or on the 
whole periphery; others become crescents, triangles, quadrangles, spindles, 
threads, etc. (see Fig. 125, a, b). 

It is customary to call anything in the plasma a platelet which is smaller 
than a red blood-cell and which does not contain hemoglobin. Buds from 
red cells lose their hemoglobin, become granular or glassy "and cannot 
then be told from platelets;" ''inner bodies " extruded from red cells, 
after undergoing certain degenerative changes, '' cannot be told from (de- 

67 Locke, Jour. Med. Research, Jan., 1902. 

68 See also Puchberger, Virchow's Arch., 1903, vol. clxxi, p. 181. 



526 CLINICAL DIAGNOSIS 

generated) platelets." This is a mistake. The term t( platelets " should 
be reserved for bodies which answer the above description especially as 
regards their peculiar bluish refractibility, their stickiness and their ten- 
dency rapidly to disintegrate. Anything floating in the plasma of the blood 
specimen fcr more than a few seconds is not a platelet no matter how much 
it may resemble it, unless some special fixing fluid had been used. 

Specimens of the platelets are best obtained by placing on the well 
cleaned tip of the finger a drop of suitable reagent, pricking the skin through 
the drop so that the blood will mix with this fluid before coming in contact 
with the air, and making a fresh preparation of this mixture. The reagents 
used are the following : Picini's fluid (mercuric bichloride 2 ; sodium chloride 
4; glycerin 26; and distilled water 226); Hayem's fluid (water 200 c.c; 
sodium chloride 1 gm. ; sodium sulphate 5 gms. ; potassium iodide solution 
[water 100; potassium iodide 5; iodine in excess] 35 c.c); Kemp's fluid 
(0.9% sodium chloride solution in 2.5% formalin); Determann's fluid 
(distilled water 160; glycerin 30; sodium chloride 1; sodium sulphate 8; 
methyl violet 0.025 parts) ; or, a 10% sodium metaphosphate solution. 

To count the platelets some estimate in a fresh specimen prepared as 
above the relative number of platelets and red blood-cells and then make 
a count of the red cells. From this data the number of platelets per cubic 
millimeter may be easily calculated. Helber 69 counted them directly, 
using a leucocyte pipette (which gave a dilution of 1:30), 10% sodium 
metaphosphate as the diluting fluid and a ruled slide similar to the ordinary 
counting-chamber, save that the thickness of the blood-film is 0.02 mm. 

The normal number of platelets per cubic millimeter has been found 
250,000 (Osier); 225,000 (Determann); 245,000 (Enden); and from 190,000 
to 260,000 (average 228,000) (Helber). The count varies in the same person 
at different hours of the day so that a change in their number must be con- 
siderable to be clinically important. In the new-born for the first few days 
the count is very low. 

It is hard to classify diseases on the basis of the platelet count but most 
agree that they are increased in all the secondary anemias but especially 
the post-hemorrhagic and that during blood regeneration they may bear 
a relation to the red blood-cells of 1:10. They are increased in chlorosis, 
but are decreased in pernicious anemia and in any severe anemia which is 
doing poorly. Their greatest increase is in spenomyelogenous leukemia 
while in lymphatic leukemia their count is very low (Pratt) . They are in- 
creased in chronic diseases with cachexia and in conditions with general 
malnutrition. 

During acute fevers of long duration they first diminish but during the 
third or fourth week as the patient begins to get weak they may increase. 
In typhoid fever a rapid diminution is considered a bad sign (Turk). In 

69 Deutsch. Arch. f. klin. Med., 1904, Bd. 81, p. 316. 



THE BLOOD 527 

short, sharp fevers there is at first a decrease, then an increase, the curve 
often resembling that of the leucocytes. The more acute, the more severe, 
the more threatening the disease and the higher the temperature, the 
fewer the platelets, so that in severe malaria and pneumonia not one may 
be found. After the temperature drops, especially if by crisis, the platelets 
may rise above normal in 24 hours and continue so for 2 to 3 days, then 
return to normal. In erysipelas and septicemia there is no preliminary 
decrease but an increase from the start. In acute articular rheumatism 
there is a great increase in these corpuscles. Four cases have been reported 
in which the platelets were totally absent: a moribund case of pneumonia, 
one of nephritis, a case of pernicious anemia and one of purpura. 

Pratt 70 found no relation to exist between the coagulation time of the 
blood and platelet count. The platelets have been shown by Wright 7l to 
arise from the peripheral portion of the protoplasm of the megakaryocytes 
or bone-marrow giant cells by a pinching-orf process. 

Previously their origin was one of the most disputed points in hematology. Schultze 
and Howell considered them fragments of broken down leucocytes; Bizzozero said they 
were independent corpuscles, a view which Osier also held; Lowit said, artefacts, while 
Hayem considered them very young red blood-cells. Then for several years it was agreed 
that they were independent corpuscles until in 1897 Arnold taught that they were frag- 
ments constricted from red blood-cells or fragments of cells which had gone to pieces. 
Maximow 72 considered them the extruded inner body of the nucleoid of red cells. Engel 
thought them remnants of the nucleus. Preisich is one of the last to insist that they 
are the extruded nuclei of the red blood-cells, hence are in constant process of formation ; 
that animals with nucleated reds have no platelets; that platelets increase as the reds 
increase, and (this point is hardest to accept) that eosinophile leucocytes are phagocytes 
which have filled themselves with platelets. 

The next work of importance is that of Deetjen 73 who, using a special agar plate 
containing sodium metaphosphate, considers that he has proved them independent 
cells, motile and nucleated. 

Wlassow was an especially severe critic of Deetjen's work. He pointed out that 
although the platelets thus treated do change their shape, yet they never show true 
ameboid motion. He, therefore, still believed that they originate in the red cells. Wlas- 
sow's argument is interesting. He made quickly a fresh specimen of blood, then ran 
under the cover-glass a drop of % concentrated ( !) mercuric chloride solution; at once the 
red cells become granular and a small refractive hyaline area appears, usually at the 
periphery of the cell. From this a bud develops which is sometimes an irregular mass 
of granules. This increases in size, may become thorny, and then separates from the 
corpuscle as a platelet. These bodies may or may not contain hemoglobin. Those 
which do, later lose their color. 

Kemp, as a result of his recent interesting work on the blood at a high altitude, is 
confident that some platelets do contain hemoglobin, and hence is in doubt as to their 
origin. 

Then came the important work of Wright, who by histological methods found an 

70 Jour, of Med. Research, Aug., .1903. 

71 Arch. f. Path. Anat., 1906, vol. 182, p. 55. 

72 Arch. f. Anatomie u. Physiologie, Anat. Abth., 1899. 
j3 Virch. Arch., May, 1901, vol. clxii. 



528 CLINICAL DIAGNOSIS 

independent origin for them in the bone-marrow, and that of Cole 74 who showed that 
a serum which will agglutinate blood platelets will not agglutinate red corpuscles. This 
certainly is strong evidence against any genetic relation between platelets and red 
blood-cells. 

CHEMISTRY OF THE BLOOD 

Hemoglobinemia is a condition of the blood characterized by the pres- 
ence of free hemoglobin in the plasma. This may be demonstrated by 
examining the spectrum of the plasma obtained by centrifugalizing the 
fresh blood, or that of the serum after the corpuscles have separated by 
clotting. Hemoglobin set free in the plasma is transformed as rapidly as 
possible in the liver to bile pigment causing hypercholia and often a definite 
jaundice, but if the destruction of red cells involves at least a sixth of their 
total number the liver is not equal to the task and hemoglobinemia results. 

Hemoglobinemia may follow a wholesale destruction of red blood-cells. 
This may, as happens in severe skin burns, take the form of fragmentation 
of the corpuscles, in which case some of the fragments may be picked up by 
the spleen (causing acute "spodogenic splenic tumor") and other internal 
organs while the hemoglobin of other cells becomes free in the plasma and, 
if in sufficient amount, causes hemoglobinemia. In other cases certain blood 
poisons lake a great many red corpuscles in the blood stream. Among these 
are the toxins of the acute infectious fevers, mentioned on page 60 1, and 
the malarial parasite. As cause of the very interesting paroxysmal 
hemoglobinuria one assumes an antecedent hemoglobinemia. This, 
however, can seldom be proved although a lowered resistance of the red 
cells to mechanical injury has been demonstrated. The best explanation 
of this condition would seem to be the following : While in the blood of 2 5% 
of all persons there is an isohemolysin capable of dissolving the red blood- 
cells, not of their own blood or of other individuals belonging to the same 
blood group with themselves (see page 588), but of certain individuals of 
other groups, patients subject to attacks of paroxysmal hemoglobinuria 
have in their plasma a hemolysin of amboceptor-complement nature which 
differs from other isohemolysins in that it is capable of dissolving the cor- 
puscles of these patients' own blood (as well as those of other individuals) 
and which needs for its activation the sudden change from a low to a high 
temperature (Moss). 

Methemoglobinemia is the condition characterized by the presence of 
methemoglobin in the circulating blood. This may be the result of the ac- 
tion of certain poisons, such as potassium chlorate, antifebrin, acetanilid, 
etc. In these cases the pigment is found both free in the plasma and 
intracellular. In certain "idiopathic" cases associated with weakness, 
cyanosis and diarrhea, the methemoglobin may be entirely intracellular. 
The presence of this pigment may be demonstrated by the spectrum of 
the blood after this has been diluted sufficiently with water. It consists of 

74 Johns Hopkins Hosp. Bull., 1907, p. 261. 



THE BLOOD 529 

the 2 bands of oxyhemoglobin and also i in the red which extends from 
X620 to X645 (shading off on the 2 sides to X615 to X650) and which 
promptly disappears on the addition of ammonium sulphide. 

The Blood in Carbon Monoxide Poisoning. — The diagnosis of severe 
cases of carbon monoxide poisoning is sometimes suggested by the macro- 
scopic appearance of the venous blood which has a slightly brighter red 
tint than normal, but the condition is proven by the spectrum of a few drops 
of the blood sufficiently diluted with water. The 2 bands of the spectrum 
of carbon monoxide hemoglobin resemble closely those of oxyhemoglobin, 
except that they are slightly nearer the violet end of the spectrum and do 
not unite to form the single band of reduced hemoglobin on the addition of 
a little ammonium sulphide. Sahli warns us not to place too high an esti- 
mate on the test. Some men are so susceptible to this gas that they show 
severe symptoms of poisoning before carbon monoxide hemoglobin can 
be demonstrated while in other cases this test will soon cease to be positive 
after the patient has breathed fresh air although the toxic symptoms may 
continue for some time later. 

Sulph-hemoglobinemia, first distinguished from methemoglobinemia 
by van der Bergh, has attracted considerable attention since the paper of 
West and Clark 75 appeared. This condition is often associated with cya- 
nosis, headache, great weakness and obstinate constipation. All sulph- 
hemoglobin is contained in the red cells, none is free in the plasma. No 
free H 2 S can be demonstrated in the blood of these patients. 

The blood spectrum of these cases shows the 2 bands of oxyhemoglobin 
and a third similar in position to that of the methemoglobin, but not quite 
so far in the red (it extends from X 610 to X625) and does not disappear but 
rather is intensified by the addition of ammonium sulphide. 

Bilirubin and Urobilin in the Blood. — For the detection of bilirubin 
and urobilin in the blood, Conner and Roper 76 suggest the following 
modification of Syllaba's Method. 

Five cubic centimeters of clear blood serum are diluted with 10 c.c. of 
distilled water, 0.5 gm. of powdered sodium sulphate added, then 1 c.c. of 
5% acetic acid; then it is coagulated by heating it briefly on a water-bath. 
It is then filtered. The color of the filtrate, usually almost colorless or 
faintly pink, does not always indicate the presence or the absence of urobilin. 
It is necessary carefully to examine all filtrates spectroscopically after the 
addition of 2 or 3 drops of Lugol's solution, using a large spectroscope 
and absorption cells from 1 to 4 cm. in depth. (A small pocket spectroscope 
will give fairly satisfactory results if the filtrates are clear.) For confirma- 
tion, the filtrate is first neutralized and then tested for the green fluor- 
escence with Schlesinger's zinc-acetate solution. The specimen is allowed to 

75 Medico-Chirurgical Transactions, vol. xc, 1907. 

76 Arch, of Int. Med., 1908, vol. ii, p. 532. 

34 



530 CLINICAL DIAGNOSIS 

stand 24 hours before it is pronounced negative. The precipitate obtained 
as above, usually very faintly yellow or white in color, is boiled for a few 
minutes on a water-bath with from 20 to 30 c.c. of 5% acid alcohol (hydro- 
chloric acid 5 parts and 95% alcohol 95 parts) and filtered. This, nitrate 
is sometimes colorless, sometimes green and sometimes yellowish pink. 
The colorless nitrates contain neither urobilin nor bilirubin; the green 
filtrates contain bilirubin and on spectroscopic examination occasionally 
show a band of urobilin; the pink filtrates contain only urobilin. 

The Quantitative Determination of Bilirubin in the Blood. — Conner and 
Roper recommend for the quantitative determination of bilirubin in the blood a slight 
modification of Gilbert's method; that is, the blood serum is diluted until Gmelin's 
test is no longer positive. This reaction is supposed to disappear when the solution of 
bilirubin reaches approximately 1 : 40,000 and the assumption is made that albumin, 
hemoglobin, indican and lutein do not interfere with the play of colors. In fluids rich 
in albumin, however, the characteristic play of colors of Gmelin's reaction is seen only 
when the bilirubin is present in relatively strong concentrations; for example, between 
1 : 3,000 and 1 : 5,000. In dilutions of from 1 : 7,000 to 1 : 11,000 one gets at the 
point of contact a distinct bluish-green ring, and in weaker solutions the blue ring becomes 
finer and has a violet rather than a green reflection, but nevertheless remains distinct 
up to a dilution of about 1 : 40,000. 

A series of dilutions of the blood serum to be tested is made, using 8 or 10 flat- 
bottomed, cylindrical glass tubes of standard size (that is, they are 4 or 5 cm. long with 
an inside diameter of exactly 1 cm. It is convenient to have a block of wood or a frame 
in which these tubes can be set in a row.) Three pipettes are necessary: One holding 
1.5 c.c. and graduated accurately to % c.c. is used for measuring the blood serum; one 
holding 2 c.c. and graduated to % c - c -» f° r measuring the diluting fluid ("artificial 
serum"); and 1 with a tapering point, for measuring approximately % c.c. of the nitric 
acid reagent. Two reagents are required. The first, the artificial serum, is made by 
mixing" well the whites of several eggs with an equal volume of 0.7% sodium-chloride 
solution and placing this on ice for 24 hours. The supernatant liquid is then decanted 
and to it is added caustic soda in the proportion of 0.3 gm. to 100 c.c. It is then allowe d 
to stand 3 or 4 days, during which time a precipitate forms which will carry down the 
coloring matters of the egg-white leaving a perfectly colorless liquid which in its fluidity , 
albumin content and alkalinity closely resembles blood serum. This should be kept 
cool. It should be freshly made at least each month. The nitric-acid reagent consists 
of 200 c.c. of pure HN0 3 , 100 c.c. of distilled water and 0.06 gm. of sodium nitrate. 

The necessary amount of blood serum for a test may be obtained from 15 to 20 c.c . 
of blood obtained by venepuncture and allowed to clot. 

Into each of 6 of the glass tubes is measured with the second pipette exactly 0.5 c.c. 
of artificial serum and then, using the first pipette, increasing amounts of blood serum 
(e-g-^Ao c.c in the first tube, %, in the second tube, % in the third, 5 / 2 o i 11 the fourth, % in 
the fifth, and *%> in the sixth tube). The contents of each tube are mixed by shaking and 
then underlaid with )i c.c. of the acid reagent. Or, the artificial serum and the blood 
serum may be mixed in a beaker and then superimposed in the proper tube over the 
nitric acid. This latter method gives a sharper end-reaction. 

After the tubes have stood for exactly 30 minutes they are examined against a white 
background in good daylight (but not in direct sunlight). The examiner's back should 
be to the light and he should hold the tubes somewhat above or below the level of his 
eyes. That tube in which the dilution of blood serum is weakest in which a faint but 
distinct blue ring can be seen, is assumed to contain a solution of 1 : 40,000 bilirubin. 



THE BLOOD 531 

The strength of bilirubin in the initial blood serum can then be readily calculated 
by the following formula: 

x i^- , in which a — the number of twentieths of cubic centimeters of serum used. 

40,000a 

Example. If the tube with the end reaction contains % c.c. of blood serum, then 
„ _ i° + S x 1 _ 3 _■ 1 



5 40,000 40,000 13,333 

The original blood serum, therefore, contained I to 13,300 of bilirubin. 

The series of tubes should be so prepared that in at least 1 tube no blue line can be 
seen. It is important that the tubes be examined just half an hour after they have been 
prepared, since the reaction tends to become stronger on standing and tubes which con- 
tain no blue line in 30 minutes may after an hour or two. 

Since the blue ring can be obtained with the undiluted serum of some normal persons 
Gilbert and Hirscher believe that normal human blood, like that of certain of the lower 
animals, contains a minute quantity of bilirubin. They found this physiologic cholemia 
to correspond to a bilirubin strength of between 1 : 28,000 and 1 : 40,000, an average 
of 1 : 36,500. 

By this test slight grades of and fluctuations in jaundice may be detected and meas- 
ured. It has been of assistance in distinguishing between appendicitis and cholecystitis. 

REACTION OF THE BLOOD 

To litmus the blood is alkaline. At least 3 5 methods have been proposed 
to determine the amount of this alkalinity but with little success since 
these methods are too inaccurate; and even though they were accurate the 
titratable alkalinity they would measure would have little value in med- 
icine. 77 By alkalinity the physical chemist means a preponderance of 
free OH-ions in the blood. According to this standard the blood is slightly 
alkaline when arterial and quite neutral when defibrinated. 

Several forms of alkalinity must be considered and that of physical 
chemistry must receive first attention. 

Pure water, the standard of neutrality, at 2o°C. contains approx- 
imately 1/10,000,000 grams of H-ions to the liter and an equivalent number 
of OH-ions. That is to say, pure water is a 1/10,000,000 normal acid and a 
1/10,000,000 normal alkali. This as expressed in logarithmic notation is 
io~ 7 N, but written for the sake of simplicity, pH 7 . If we have less than 
1/10,000,000 gram of hydrogen-ions in one liter the solution is less acid than 
water, that is, it is alkaline. That is, pH 8 means actually 1/10,000,000 N 
alkaline. The higher the exponent, the more alkaline, that is, the less acid, 
the solution. 

pH 1 = N/10 acid 

pH 6 = N/i, 000,000 acid 

pH 7 = Neutrality 

pH 8 = N/ 1, 000, 000 alkali 

pH 14 = N/10 alkali 

77 Strouse, Johns Hopkins Hospital Bulletin, Vol. 19, p. 139, May, 1908. 77a In the 
preparation of the following sections on the chemistry of the blood we have used freely 
Gradwohl and Blaivas' "Newer Methods of Blood and Urine Chemistry". 



532 CLINICAL DIAGNOSIS 

The reaction of the blood serum always lies between pH 7 and pH 8 . 
The neutral point, pH 7 , is reached only in severe uncompensated acidosis, 
while a reaction of pH 8 is attained perhaps only after the administration 
of large doses of alkalies. 

Since the electro-chemical methods of determining the reaction of the 
blood are too complicated for clinical work the attempt has been made 
to find a color indicator which would show definite progressive and con- 
stant color changes for each degree of the H-ion concentration. 

Marriot, Levy, and Rowntree's Method for the Determination 
of the hydrogen-ion concentration of the blood. 78 

Levy, Rowntree and Marriott used phenolsulphonphthalein which 
exhibits definite variations in color corresponding to very minute differences 
in hydrogen-ion concentration between pH 6-4 and pH 8,4 . They prepared 
a series of standard solutions of known pH colored by this indicator, and 
then added an equal amount of this indicator to the solution to be 
tested and determined which of the colors of the standard series it most 
closely matched. 

Since both blood and serum possess color and since proteins interfere 
with the colors of many indicators and it is impossible to apply the method 
directly to the blood, they dialyzed the serum or plasma of the whole, or 
of the denbrinated, blood in collodion sacks for 5 minutes, added the indi- 
cator to the dialysate and matched the resulting color to the standard 
series. This method while still in use has not proven accurate enough to 
win recognition. 

The method more generally in use is : 

Van Slyke's Method for the Determination of the Carbon Di- 
oxide Combining Power of the Blood Plasma. Twenty-five cubic 
centimeters of blood drawn from the vein are allowed to run from the needle 
directly into a small chemical bottle which contains 10 drops of a 20% so- 
lution of potassium oxalate. (The oxalate used should be dried in the oven 
over night at ioo°C. before the solution is made.) The bottle is quickly 
closed and shaken vigorously until perfect fluidity of the blood has been 
obtained. It is then at once centrifugalized, the clear serum pipetted off 
and placed in a separating funnel of about 300 c.c. capacity. While 
hemolysis should be avoided as much as possible by immediate centrifu- 
galization yet slight hemolysis will not appreciably affect the results. 
To determine its alkaline reserve, the plasma is saturated with carbon 
dioxide at alveolar tension by the operator who blows vigorously through 
a bottle containing glass beads into the separating funnel, as shown in Fig. 
126. If one were to blow directly into the funnel enough moisture would 
condense on its walls to dilute appreciably the plasma. The funnel should 

be closed at the stop-cock S and with the stopper T just before the stream of 

i_ , , _ __ 

78 Levy, Rowntree, and Marriott: Arch, of Int. Med., 1915, vol. xvi, p. 389. 



THE BLOOD 533 

breath stops and shaken for i minute in such a manner that the plasma will 
be distributed as completely as possible on its walls. After this shaking 
more alveolar air is blown through the beads into the funnel and the 
shaking repeated for i minute. 

The C0 2 apparatus (Fig. 127) is held in a strong clamp W, which is 
lined with rubber, and the lower stop-cock is supported by an iron rod 
which also is covered with soft rubber tubing. The apparatus, which should 
be thoroughly cleaned before the determination is started, is completely 
filled with mercury. Care should be taken that capillaries A and F, which 
are above the upper stop-cock, also are filled with mercury and that there 
are no air bubbles anywhere within the apparatus. This can be tested by 
lowering and raising the bulb, thus allowing the mercury to run down and 
up until there is a not a single air bubble in the apparatus. Six dropping 
bottles, which contain the following solutions, should be at hand: 

1. Distilled water. 2. Phenolphthalein (1% in 95% alcohol). 3. Nor- 
mal ammonium hydroxide. 4. Capry lie alcohol. 5. Normal sulphuric acid. 
6. Mercury. 

The mercury leveling bulb H should be hung by a wire on the extension 
N about on the level with the lower cock /. One drop of phenolphthalein 
and a drop or two of the ammonium hydroxide are then placed in the upper 
cup B. About }4 c.c. of distilled water is then added and then all drawn 
off except about 2 drops of the alkaline solution. 

One cubic centimeter of the saturated plasma is now introduced into 
the cup B and allowed to flow under the alkaline solution so that none of 
the carbon dioxide will escape. The stop-cock C is then turned so that the 
tubes E and Z are connected and the plasma allowed to run in until the 
capillary Fis exactly rilled. Distilled water, 0.5 c.c, is now measured into 
cup B and allowed to run down to the capillary F. This is repeated , care 
being taken that no air enters the apparatus with the liquid. Next 1 drop 
of capry lie alcohol is admitted into the capillary F to prevent foaming, and 
then about 1.5 c.c. of the sulphuric acid poured into the cup. Enough of 
the acid is next admitted into the apparatus, carrying the capry lie alcohol 
along with it, so that the total volume reaches exactly to the 2.5 c.c. mark. 
The excess of sulphuric acid is now drawn off. A few drops of mercury 
are now placed in the cup B and allowed to now down to the capillary F 
in order to seal this and make it capable of holding an absolute vacuum. 
During this whole operation the lower stop-cock J should remain open and, 
when the apparatus is set up, it should be in such adjustment that if the 
wire I which is connected to the bulb H is lowered to the hook the mercury 
will run to the middle of the small bulb just above the fork but not into 
the fork Y. The wire I is now placed on the hook and the mercury allowed 
to fall until its meniscus has dropped to the 50 c.c. mark on the apparatus. 
This is controlled by the stop-cock /. The bubbles of C0 2 will now be 
seen escaping. 



534 CLINICAL DIAGNOSIS 

In order to extract the carbon dioxide completely the apparatus is re- 
moved from the clamp and shaken by turning it upside down about a 
dozen times. (The thumb should be placed over cup B so as not to lose any 
of the mercury.) The apparatus is then replaced, the mercury leveling 
bulb H still being at the low level 0, and the solution allowed to flow into 
the small bulb below the lower stop-cock (right side). The solution is 
drained out of the portion of the apparatus above the stop-cock / as com- 
pletely as possible, but without removing any of the gas (the last drop being 
allowed to remain above). The mercury bulb H is now raised in the left 
hand and with the right hand the lower stop-cock J is immediately turned 
so that the mercury is admitted to the upper part of the apparatus through 
the left-hand entrance of the stop-cock without readmitting the watery 
solution. The bulb H is now held beside the apparatus so that its mercury 
level corresponds to that in the apparatus and the gas in the latter is under 
atmospheric pressure. A few hundredths of a cubic centimeter of water 
will float on the mercury in the apparatus but this may be disregarded in 
leveling. The calculation of the result in terms of volume percentage of 
carbon dioxide bound as carbonate by the plasma is quite complicated and 
so Van Slyke recommends that we consequently use the direct reading from 
the apparatus minus 0.12. 

The plasma of normal adults yields from 0.65C.C. to 0.90 c.c. of gas which 
is the direct reading on the apparatus. If 0.12 is subtracted, the normal 
figures would be from 53 to 78 in terms of volume per cent, of carbon dioxide 
chemically bound by the plasma. Figures lower than 50% in adults would 
indicate acidosis. The results thus obtained give approximately (within 2 
or 3%) the volume per cent, of carbon dioxide bound by the plasma. 

Example. — The reading on the Van Slyke apparatus is, e. g., 0.74. This 
minus 0.12 equals 0.62% of carbon dioxide bound by 1 c.c. of plasma, 
or, for 100 c.c, 62% which is normal. 

The Determination of the Alkali Reserve of the Blood Plasma. — 
Marriott 79 has recently published a method of determining the hydrogen- 
ion concentration of the dialysate of blood serum after the removal of the 
carbon dioxide which is more accurate and helpful than the preceding test, 
since it serves for the detection and accurate quantitative estimation of the 
degree of an acidosis. 

The apparatus required includes: a set of tubes containing standard 
phosphate mixtures; a solution of phenolsulphonphthalein in 0.8% sodium 
chloride; collodion sacks; a pipette to measure 0.5 c.c; small test tubes for 
dialyzing and aerating; an atomizer bulb; a glass tube or pipette drawn out 
to a fine capillary point and a box for comparing colors. 

A 1/15 molecular acid, or primary, potassium phosphate is made by dis- 
solving 9.078 gms. of the pure recrystallized salt (KH2PO4) in freshly 

79 Arch. Int. Med., June, 1916, vol. xvii, p. 840. 



THE BLOOD 



535 



distilled water, exactly 200 c.c. of a 0.01% solution of phenolphthalein is 
added and the solution made up to 1 liter. 

One-fifteenth molecular alkaline, or secondary, sodium phosphate solu- 
tion is made by exposing the pure recrystallized salt (Na 2 HP04.i2H 2 0) 
protected from dust to the air for from 10 days to 2 weeks, (Ten molecules 
of water of crystallization will be given off yielding a salt of the formula 
Na 2 HPO 4 . 2H 2 0) , dissolving 11.876 gms . of this in freshly distilled water, add- 
ing exactly 200 c.c. of a 0.017% solution of phenolphthalein and making 
the solution up to 1 liter. The solution should give a deep rose-red color 
with phenolphthalein. If only a faint pink color is obtained the salt is not 
sufficiently pure. The exact amount of indicator is immaterial, provided 
the same amount is added to each of the phosphate solutions. A small 
crystal of thymol also is added to each to prevent the growth of moulds. 
These solutions should be preserved in Jena or Non-sol glass vessels. 
These solutions are mixed in the proportions indicated below to obtain the 
desired pH. 



pH 

Primary sod. phos., c 
Secondary sod. phos. 



7.0 


7.2 


74 


7.6 


7.8 


8.0 


8.2 


8.4 


37-0 


27.0 


19.0 


13.2 


8.8 


5.6 


3-2 


2.0 


63.0 


73-o 


81.0 


86.8 


91.2 


944 


96.8 


98.0 



8.6 

1.0 

99.0 



These solutions are measured into small test tubes, approximately 100 
mm. long by 8 mm. internal diameter, made of glass that does not readily 
give off alkali and these stoppered or sealed. They should be kept in a 
dark place when not in use. Under these conditions, the solutions will 
retain their colors for long periods of time. 

The salt solution is made up by dissolving 8 gms. of chemically pure 
sodium chloride in distilled water, adding 220 c.c. of 0.017% phenolsul- 
phonphthalein solution and making the whole up to 1 liter with distilled 
water. The intention is that this solution shall have just the same strength 
of indicator as the 2 phosphate solutions, but since a certain amount of that 
in the salt solution is lost during the dialysis by passing into the sack it is 
made to contain an initial excess of 10%. To test this solution for free 
alkali and acids other than carbonic, a little of it is boiled (or aerated with a 
current of air that has been freed from carbon dioxide by passing through a 
strong solution of sodium hydroxide) for a minute or so in a test tube of hard 
Jena glass (which will give off no alkali) in order to expel the carbonic acid. 
It is then cooled quickly under the tap and compared with the phosphate 
standards. Its reaction should be 7.0. If not, it may be corrected by the 
addition to the whole solution of a few drops of very dilute acid or alkali. 
This solution should be kept in a vessel of Jena or Non-sol glass, or in a 
vessel of ordinary glass that has been well paraffined on the inside. 

The determination must be carried out in a room free from acid and 
ammonia fumes. While serum, oxalated plasma, or blood may be used, 



536 CLINICAL DIAGNOSIS 

serum is to be preferred since the addition of oxalate, unless exactly neutral, 
will introduce a source of error. The blood should be collected in a small 
tube and the serum separated as quickly as possible, preferably by centrif- 
uging, otherwise there will be a passage of some of the alkali of the plasma 
to the cells following the loss of some carbon dioxide. Hemolysis must 
be avoided. 

Exactly 0.5 c.c.of serum is pipetted into one of the small collodion sacks, 
which has previously been washed inside and out with the salt solution. 
(In washing the sack, no part but the top edge should be touched with the 
ringers. The sack is emptied by tipping it with a clean glass rod or with 
a microscopic slide. Sacks may be used more than once, providing they are 
thoroughly washed with salt solution after each test.) The sack is then 
lowered into a small test tube, approximately 8 mm. internal diameter and 
50 mm. long, containing 2 c.c. of the indicator salt solution. The level of 
the fluid on the outside of the sack should at least be as high as that on the 
inside. At the end of 7 minutes the sack is removed and the dialysate 
transferred to a clean test tube from 100 to 140 mm. long and having the 
same diameter as the tubes containing the phosphate standards. A rapid 
current of air is now bubbled through the solution in order to remove the 
carbon dioxide. This may be done with an atomizer bulb connected with 
a narrow glass tube drawn out to a capillary point. (Troublesome foaming 
may be prevented by adding a drop of octyl alcohol or toluol.) The blowing 
is continued for 3 minutes and then the color in the tube is compared with 
that of the standard phosphate tubes. The reading is a measure of the 
reserve alkalinity which is expressed as RpH. 

In the case of every normal person on a general mixed diet the RpH of 
the serum was found tobe8.5io.05. A normal adult's serum drawn after 
a fast of 16 hours gave a reading of 8.35. The serum of normal infants 
under 1 year of age frequently gives values as low as 8.3. This accords 
well with the observation that the carbon dioxide tension in the alveolar air 
is lower, that the combined carbon dioxide of the ■ plasma is less and that 
the ammonia coefficient in the urine is often higher in infants than in 
adults. This slight but normal acidosis might well be the result of the 
more active metabolism of infants, leading to a proportionately greater 
production of acids, although the fact that infants' blood is usually obtained 
for this test by cupping should not be overlooked. 

In all the cases of acidosis studied (including cases of nephritis and 
diabetes in adults, and nephritis, recurrent and idiopathic acetonemia and 
severe diarrheas in children. The diarrheal cases were of the type described 
by Howland and Marriott) the RpH of the serum showed deviations from 
the normal. The more severe the acidosis (as judged by other methods) 
the lower the RpH. Especially striking was the parallelism, between al- 
veolar carbon dioxide tension and the RpH in all cases without any disturb- 
ance of the respiratory center or lesion of the pulmonary epithelium. 



THE BLOOD 537 

The results obtained by Van Slyke's method of determining the carbon 
dioxide combining power of the blood plasma (see page 533) were in a. 
general way proportional to the RpH of the serum. In cases of alkali 
starvation the RpH gave information as to the probable amount of alkali 
needed to replenish the reserve while a determination following the 
administration of alkali showed whether or not the amount given 
was sufficient. 

The values obtained for the RpH of the serum may, in the light of 
Marriott's experience, be interpreted as follows: Values for RpH of from 
8.4 to 8.55 correspond to alveolar carbon dioxide tensions of from 38 to 
45 mm. and are to be considered as normal values for adults. Values 
between 8.0 and 8.3 correspond to alveolar carbon dioxide tensions of from 
28 to 35 mm. and indicate a moderate degree of acidosis. 

When the value for RpH is as low as 7.7, corresponding to an alveolar 
carbon dioxide tension of 20 mm., the individual is in imminent danger. 
During coma an RpH as low as 7.3, corresponding to an alveolar air 
tension of 11 mm. was observed. In infants under one year of age a value 
for RpH of 8. 3, corresponding to 35 mm. tension in the alveolar air, is not 
to be considered abnormal. 

It has been Marriott's experience that unless the RpH of the serum is 
below 7.9 the acidosis may be successfully combated by dietetic regulation 
or by the administration of alkali by mouth. When the RpH of the serum 
falls below 7.9 intravenous administration of alkali is usually indicated. 

Lowy Method. — Lowy's method is still the best for determining the 
total alkalinity of the blood. Into a special flask, on the neck of which 
are marks indicating 45 and 50 c.c, are measured 45 c.c. of 0.25 % am- 
monium oxalate. To this are added 5 c.c. of blood. The blood is therefore 
laked and will not coagulate, hence the alkalinity determined is that of 
the total blood. This is then titrated with 0.04N tartaric acid using 
lacmoid paper saturated with magnesium sulphate as indicator. By this 
method it has been determined that 100 c.c. of normal blood contains from 
400 to 600 mgms. of NaOH. 

Salkowski's Method. — Salkowski's method is certainly simple. 
A known amount of ammonium sulphate is mixed with the blood and the 
ammonia set free determined by Schlosing's method (see page 123). 

Under a bell-jar is' placed a dish containing 20 gms. of finely pulverized 
ammonium sulphate dissolved in 20 c.c. of water. In a receptacle above 
this is 10 c.c. of 0.25 N H2SO4. Into the lower dish is poured 10 c.c. of the 
blood to be examined (the measuring-glass used for the blood is first washed 
in 1% sodium oxalate solution to prevent coagulation). The blood is mixed 
with the ammonium sulphate solution and the apparatus covered at once 
with the bell- jar. In 5 or 6 days all of the ammonia set free from the am- 
monium sulphate will have been taken up by the sulphuric acid and its 
amount may be determined by titrating this. By this method the alkalinity 



538 CLINICAL DIAGNOSIS 

for men has been found to be from 350 to 400 mgms. and for women from 
300 to 350 mgms. of NaOH per 100 c.c. of blood. 

Sellard's Method (Qualitative) for Determining the Titratable 
Alkalinity of the Blood. 80 One cubic centimeter of serum is shaken 
up thoroughly with 25 c.c. of absolute alcohol and filtered into an evaporat- 
ing dish. Perfectly dry apparatus should be used. Without washing the 
precipitate 3 or 4 drops of an 0.5% solution in absolute alcohol of phenol- 
phthalein are added to the filtrate and this evaporated to dryness on a 
steam bath (at not over ioo°C). 

In the case of normal serum the residue is continuously red. In the 
presence of an acidosis the residue is colorless or pink but on the addition of 
1 drop of water becomes an intense red. In case of an extreme acidosis it 
remains colorless even after the addition of water. 

To prove the neutrality of the absolute alcohol 0.1 c.c. of 0.005N NaOH 
is added to 25 c.c. of the alcohol, also 2 or 3 drops of phenolphthalein as 
indicator, and the specimen evaporated to dryness. The residue should 
be distinctly pink. 

CHLORIDES OF BLOOD 

To determine the chlorides of the blood 3 c.c. of the blood to be examined 
are measured with a pipette into a 50 c.c. graduated centrifuge tube, 15 c.c. 
of 0.0 1 N acetic acid added and the specimen diluted to 30 c.c. with distilled 
water. The tube is then placed in a beaker of boiling watei to bring about 
the coagulation of the protein, care being taken that the contents of the 
tube aie agitated occasionally with a stirring rod. After the protein has 
coagulated the tube is cooled, againmade up to volume (30 c.c.) and centrif- 
uged. The slightly colored supernatant fluid is now poured into a dry 
centrifuge tube, about 6 drops of a strong solution of colloidal iron added 
and the tube placed in a beaker of hot water for a few minutes. This will 
completely precipitate all the protein. The specimen is then centrifugalized 
or filtered once more and 10 c.c. (equivalent of 1 c.c. of blood) of the clear 
fluid poured into a 2 5 c.c. volumetric flask together with 10 c.c. of the stand- 
ard silver nitrate solution 81 and 1 c.c. of the ferric alum indicator 82 added. 

The fluid is now made up to volume and shaken thoroughly, centrif- 
ugalized in a large (50 c.c.) centrifuge tube and the clear supernatant 
fluid decanted. One then titrates 20 c.c. of this fluid, which is the equivalent 
of 0.8 c.c. of blood, with a standard ammonium thiocyanate solution 83 of 

80 Bull, of the Johns Hopkins Hosp., Apr., 1914, vol. xxv, p. 101. 

81 This standard is prepared by dissolving 2.906 gms. of silver nitrate in distilled 
water and making up to 1 liter, therefore 1 c.c. =0.001 gm. of sodium chloride. 

82 This indicator is made by dissolving 100 gms. of crystalline ferric ammonium 
sulphate in 100 c.c. of 25% nitric acid. 

83 Standard ammonium thiocyanate solution is prepared by dissolving _ 1.3 gms. of 
ammonium thiocyanate in 800 c.c. of water, titrating against the above silver nitrate 
standard and ascertaining the amount of water which must be added to the_ solution 
to make each 1 c.c. of it equivalent to 1 c.c. of the standard silver nitrate solution; that 
is, to indicate 0.001 gm. of sodium chloride. 






THE BLOOD 539 

the same strength as the silver nitrate, until a distinct yellow color shows 
throughout the mixture. The titration result, divided by 0.8, subtracted 
from 10, to obtain the silver nitrate used, and multiplied by .001 and again 
multiplied by 100, gives the percentage in the blood of chlorides as sodium 
chloride. 

Example. — Suppose the reading on burette is 3.2 c.c. This divided 
by 0.8 = 4: 10 — 4 = 6:6X-ooiXioo = 0.6, the percentage of chlorides (as 
NaCl) in this specimen of blood. 

NITROGENOUS BODIES OF THE BLOOD 

Total Nitrogen. — To determine the total nitrogen of the blood exactly 
1 c.c of the specimen to be examined is measured into a long-necked Jena 
glass Kjeldahl flask, 20 c.c. of concentrated sulphuric acid and about 
0.2 gm. of copper sulphate added and the mixture boiled in the digestion 
rack for about 1 hour after it has become colorless. The flask is then cooled 
and the contents diluted with about 200 c.c. of ammonia-free water. Then 
one adds about 40 c.c. of a saturated sodium hydroxide solution, that is, 
a little more than is necessary to neutralize the sulphuric acid, and a little 
coarse pumice stone or a few pieces of granulated zinc to prevent thumping 
and a small piece of paraffin to lessen the tendency to froth. The flask 
is then connected by means of a safety tube with a condenser so arranged 
that the delivery tube passes into a vessel containing a known volume (the 
volume used depending upon the nitrogen contents of the blood) of 0.1N 
H 2 S0 4 to which has been added a few drops of congo red. (Congo red 0.5 
gm. dissolved in a mixture of 90 c.c. of distilled water and 10 c.c. of 95% 
alcohol.) The end of the delivery tube must reach beneath the surface of 
the fluid and the tube should be of a large enough caliber to avoid the suck- 
ing back of the fluid. The contents of the distillation flask should be mixed 
very thoroughly by shaking (or rotating) and the mixture then distilled 
until about % of the solution has passed over. The partly neutralized 
0.1N H 2 S0 4 is now titrated against 0.1N NaOH. In this way one deter- 
mines the amount of 0.1N H 2 S0 4 neutralized by the ammonia which has 
distilled over. 1 c.c. of o.i7VH 2 S04 is the equivalent of 0.0014 gm. 
nitrogen. This multiplied by 100 is the amount of N in 100 c.c. of blood. 

Folin-Farmer Microchemical Method. — Exactly 1 c.c. of the blood 
to be examined is pipetted into a 25 c.c. volumetric flask, diluted with dis- 
tilled water up to the 25 c.c. mark and well mixed. One cubic centimeter 
of the diluted blood is pipetted into a test tube of such a size that it will 
slip into the aeration apparatus. From 0.1 to 0.3 gm. of potassium sulphate, 
a drop of 10% copper sulphate solution and 1 c.c. of concentrated sulphuric 
acid are added in the order named and the digestion carried out as in the 
determination of non-protein nitrogen (see page 540). The result obtained 
above is for % 5 c.c. of blood. 

Example. — Suppose the dilution was to 50 and the reading 75 which 



540 CLINICAL DIAGNOSIS 

on the scale would indicate 0.56 mgm. had the dilution been to 100. There- 
fore 0.56-^2 = 0.28 mgm., in 2 c.c. of blood or i4mgms. in 100 c.c. 

To calculate the amount of urea which a given amount of nitrogen 
would represent the value of the latter is multiplied by the factor 2.14. 

Non-proteid Nitrogen of the Blood. — The non-proteid nitrogenous 
products of catabolism which are constituents of the blood and which may 
accumulate in cases of renal insufficiency are: 84 Urea, formed largely in 
the liver from the ammonia resulting from the deaminization of amido 
acids set free in digestion but not of immediate use to the animal organiza- 
tion; uric acid, a product of the enzymatic transformation of the amino- 
and oxypurins, in which various glands of the body participate; and crea- 
tinin, which would appear to be formed in the muscle tissue from 
creatin. Under normal conditions the percentage relationship of these 
bodies to total non-proteid nitrogen is: urea 50%, creatinin 2%, uric 
acid 2%, ammonia 0.3% and undetermined nitrogen 4.6%. In the urine 
these figures are: urea 85%, creatinin 5%, uric acid 1.5%, ammonia 4% 
and undetermined nitrogen 4.5%. Judged by their comparative composi- 
tion the kidney concentrates the creatinin 100 times, the urea 80 times 
and the uric acid only 20 times. Creatinin is therefore of these 3 bodies 
the most readily eliminated, urea next and uric acid least. 

To determine the non-proteid nitrogen 5 c.c. of the blood to be examined 
are measured into a 50 c.c. volumetric flask containing about 35 c.c. of 
2.5% trichloracetic acid and the volume then made up to 50 c.c. with this 
same acid. The flask is then shaken vigorously. At the end of 30 minutes 
(or as soon after as convenient) the specimen is filtered through a dry filter. 
To the nitrate are added about 2 gnus, of kaolin and it is shaken 
vigorously. The mixture is then allowed to stand for from 5 to 10 
minutes and the nitration repeated. This filtrate should be quite colorless. 
One next pipettes 10 c.c. of the filtrate (the equivalent of 1 c.c. of blood) 
into a test tube about 200 mm. long and of a sufficient diameter to slip 
into a 100 c.c. graduated cylinder (see Fig. 27). From 0.1 to 0.3 gm. of 
potassium sulphate, a drop of 10% copper sulphate and 1 c.c. of concen- 
trated sulphuric acid are added in the order named (these reagents should 
be of the highest purity). This is then boiled over a microburner, at first 
gently, until a dark brown color appears. Gradwohl and Blaivas recommend 
that the solution then be allowed to cool and a drop of peroxide of hydrogen 
added. If the mixture does not clear it is heated gently over the micro- 
burner and, if that is not sufficient, this process is repeated. The tube is 
now allowed to cool and about 5 or 6 c.c. cf distilled water added. 

As a means of removing the fumes, the suction pump is connected by a 
2-hole stopper to a large bottle (Fig 128) containing a solution of sodium 
hydroxide. The tube B should be attached to a funnel held over the mouth 

84 Quoted from Myers, The Jour, of Lab. and Clin. Med., Apr., 1920, v 3 p. 418. 



THE BLOOD 541 

of the test tube D. After a few determinations have been made, it is well 
to wash the funnel to remove any acid which may have condensed upon it. 

Aeration is carried out exactly in the manner for urea, the only difference 
being that saturated sodium hydroxide is used instead of saturated sodium 
carbonate. The same table is used for the calculations. 

Blood Urea. — Marked urea retention occurs in a wide variety of con- 
ditions: in the terminal stages of chronic interstitial nephritis, in some 
cases of acute nephritis (but it is not at all marked in parenchymatous 
nephritis), in bichloride and in lead poisoning, in congenital cystic kidney, 
malignancy, pneumonia, intestinal obstruction and sometimes in syphilis 
and cardiac conditions. A normal urea with a high uric acid content in the 
blood is of value in the diagnosis of gout. In eclampsia the blood urea is but 
slightly elevated if at all, while in normal pregnancy the figures are low. 

The blood urea will vary from the normal only late in a case of renal 
insufficiency, much later than creatinin and uric acid, and its determination 
is of little value in diagnosis but is of value in treatment. If increased 
blood urea may be reduced by increasing the fluid intake and decreasing 
the proteid of the diet. 

Case 9559, a woman 45 years old, was admitted on May 3, 1920 Two weeks previ- 
ous to this time her blood creatinin was 2.4 mgms. per 100 c.c. and her phenolsulpho- 
nephthalein output. 41% in 2 hours. On May 5 the blood creatinin was 3.29 mgms., 
the blood urea 35.04 mgms. per 100 c.c, and the phenolsulphonephthalein output 25%. 
On May 13 the blood creatinin was 3.97 mgms. and the blood urea 30 mgms. On 
May 22 the blood creatinin was 4. 11 mgms. and the blood urea 145.2 mgms. On May 
25, the blood creatinin was 14.12 mgms. and blood urea 350.4 mgms. per 100 c.c. Then 
came the first convulsion. She died May 28, 1920, in uremia. 

Urea. — -For the determination of urea 2 c.c. of the blood to be examined 
are measured with an Ostwald pipette into a test tube which already con- 
tains 2 c.c. of distilled water and 0.1 gm. of urease. This tube should be of 
such size that it will readily slip into a 100 c.c. graduated cylinder. This 
is then incubated for }4 hour in a beaker of water at 5o°C. At the end of 
this time 2 drops of caprylic alcohol or 1 c.c. of amy lie alcohol aie added to 
prevent foaming in the subsequent aeration. The urease converts the urea 
into ammonium carbonate. The ammonia is then liberated by an excess 
of sodium carbonate and carried over by suction into hydrochloric acid 
where it forms ammonium chloride. The ammonia may now be determined 
colorimetrically by the use of Nessler's reagent. The apparatus is that 
used for the determination of urea in the urine (see page 113) 

Into cylinder 1 are poured 20 c.c. of distilled water and 2 to 3 drops of 
10% HC1. This cylinder is now closed and cylinder 2 opened! To the 
test tube containing the digested blood an equal volume of saturated sodium 
carbonate is allowed to run slowly down under the blood, this placed at 
once in the cylinder and this immediately closed , with the long tube dipping 
practically to the bottom of the fluid in the test tube and the connection 



542 



CLINICAL DIAGNOSIS 



carefully sealed. The suction by a Chapman pump is slow for about 5 
minutes and then gradually increased to as much as the apparatus will stand. 
The aeration is kept up for from 30 to 45 minutes. At the end of this time 
the pump is disconnected, the rubber stopper from cylinder 1 removed and 
the tube washed off with 2 or 3 c.c of distilled water. 

The color test is made as follows. Into a 50 c.c. volumetric flask is 
pipetted 5 c.c. of ammonium sulphate solution containing 1.0 mgm. of 
nitrogen per 5 c.c. of the solution (this is the standard solution) (see page 
109) and then 25 c.c. of distilled water and 20 c.c. of Nessler's solution 
(see page 109) diluted 1 to 5 added. 

To cylinder 1, containing the ammonia in the form of ammonium 
chloride, add from 10 to 20 c.c. of diluted Nessler's solution (1 to 5), de- 
pending upon the depth of color, and then dilute to 50 c.c, 100 c.c, etc., 
according to the color. The colorimetric reading should be made at 
once and computed from the following table : 

TABLE IV* 

Estimation of Nitrogen with the Hellige Colorimeter 





Nitrogen 




Nitrogen 




Nitrogen 


Colorimetric 


mgms. per 


Colorimetric 


mgms. per 


Colorimetric 


mgms. per 


reading 


dilution of 


reading 


dilution of 


reading 


dilution of 




100 c.c. 




100 c.c. 




100 c.c. 


20 


i-73 


40 


131 


60 


O.89 


21 


1. 71 


41 


I.29 


6l 


O.87 


22 


1.69 


42 


I.27 


62 


O.85 


23 


1.67 


43 


I.25 


63 


O.83 


24 


1.65 


44 


I.23 


64 


0.8l 


25 


1.62 


45 


1.20 


65 


O.78 


26 


1.60 


46 


1. 18 


66 


O.76 


27 


1.58 


47 


1. 16 


67 


O.74 


28 


i-56 


48 


1. 14 


68 


O.72 


29 


i-54 


49 


1. 12 


69 


O.7O 


30 


1.52 


50 


1. 10 


70 


O.67 


31 


1.50 


5i 


1.08 


7i 


O.65 


32 


1.48 


52 


1.06 


7^ 


O.63 


33 


1.46 


53 


I.04 


73 


0.6l 


34 


1.44 


54 


I.02 


74 


o-59 


35 


1.41 


55 


O.99 


75 


0.56 


36 


i-39 


56 


O.97 


76 


0-54 


37 


i-37 


57 


0.95 


77 


0.52 


38 


i-35 


58 


0-93 


78 


0.50 


39 


i-33 


59 


O.9I 


79 


0.48 



'Myers and Fine: Table copied from Gradwohl and Blaivas. 



Ammonia. — In the determination of urea any preformed ammonia 
would be determined at the same time. As a correction for the urea de- 
termination necessary in some cases, and also if the ammonia is to be 
determinated separately, a specimen of blood is examined exactly as above 
except that no urease is used. The blood should be examined immediately 



THE BLOOD 543 

after it is drawn from the vein since on standing the ammonia begins at 
once to increase. 

Under normal conditions 85 the ammonia of the blood varies from 0.4 
t o o . 7 5 mgms . , never more than 1 mgm . , per 1 00 c . c . An amount of 3 m gms . 
or over per 100 c.c. may certainly be considered abnormal. Normally, 
diet has no appreciable influence on the ammonia although for the 
sake of uniformity the patients are placed for at least 1 day on a diet con- 
sisting of at least 32 ounces each of milk and albumin water per 24 hours. 

McNeil and Levy found that an increased ammonia content of the 
blood could not in all cases be explained by the presence of an acidosis, nor 
was it nearly as common in cases with advanced hepatic disease as one 
would expect. In one man with periodic attacks of nausea and vomiting 
an ammonia content of 23 mgms. per 100 c.c. of blood was found. The 
ammonia was not increased in the blood of four patients with uremia, but 
was in several cases of eclampsia. 

Uric Acid in the Blood. — Under normal conditions the blood contains 
from 1 to 3.5 (average 2) mgms. of uric acid per 100 c.c. This is increased 
physiologically during the first 3 or 4 days of life and pathologically in 
gout, lead and mercuric bichloride poisoning, malignancy, acute infections 
especially pneumonia, in leukemia and in chronic insterstitial (not paren- 
chymatous) nephritis. These increases may be due to increased production, 
as in leukemia, or to decreased elimination, as in nephritis. In very early 
stages of interstitial nephritis, before any concentration in the blood of urea 
or creatinin can be demonstrated, that of uric acid may be very definite, 
which makes this an early diagnostic sign of this disease and perhaps the 
most delicate index of renal function we have. 86 Later, after the renal 
insufficiency has become marked, the uric acid content may reach 10, 15 and 
in one case just before death 27 mgms. per 100 c.c. of blood. In gout the 
uric acid of the blood is invariably increased, ranging from 3.5 to 5.5 mgms. 
and even 9 mgms. per 100 c.c. of blood. (The reader will have in mind the 
common, perhaps invariable, association of chronic interstitial nephritis 
with gout. Indeed the old term applied to the small contracted kidney 
was " gouty kidney." It is an old idea now again advanced that gout 
is merely a form of chronic interstitial nephritis with conspicuous 
j oint complications . 87 ) 

In other forms of arthritis, excepting those with a definite renal complica- 
tion the uric acid varied from 1.6 to 3.6 mgms. (the majority below 3 
mgms.) per 100 c.c. of blood. 

For the estimation of uric acid in the blood 88 10 c.c. of the specimen 
to be examined are measured into a casserole of at least 375 c.c. capacity 

85 McNeil and Levy, Jour, of Lab. and Clin. Med., 191 7, iii, 18. 

86 See Myers, Fine and Lough, Arch, of Int. Med., 19 16, vol. xvii, p. 570. 

87 Jour. A. M. A., 1916, lxvi, 2051. 

88 Benedict's method as given by Gradwohl and Blaivas. 



544 CLINICAL DIAGNOSIS 

and 50 c.c of 0.0 iN acetic acid added. 89 The casserole is placed on a 
water-bath and heated until coagulation takes place, then over a free 
flame until the contents come to a boil, stirring continuously. 
About 1 spoonful (4 c.c.) of alumina cream 90 is now added and the 
boiling continued for one minute, stirring continuously. The speci- 
men is now filtered and the coagulum on the filter paper washed back into 
the casserole with about 100 c.c. of hot distilled water. This mixture in the 
casserole is next heated to the boiling point over a free flame and then fil- 
tered . The combined filtrates are evaporated by boiling, then slowly over 
a free flame, until their volume has been reduced to about -500 c.c, then in 
the water bath until it is evaporated down to 1 or 2 c.c. This is then trans- 
ferred to a conical centrifuge tube of 15 c.c. capacity, washing the casserole 
with 2 or 3 portions of hot water but keeping the final volume below 10 c.c. 
When this has cooled, 15 drops of ammoniacal-silver-magnesium mixture 
(see page 119) are added, the tube is shaken and placed in a refrigerator for 
about 1 5 minutes (to allow the precipitation of purine) . It is then centrif- 
ugalized for from 3 to 5 minutes and the supernatant fluid then poured off 
by inverting the tube and wiping its lip with filter paper. The ammonia of 
the sediment is then removed by suction. This is accomplished by attaching 
the centrifuge tube to the rubber tubing of the Chapman pump. 

We are now ready for the development of color and for the reading. 
The beginner should work as fast as possible as the color may fade or tur- 
bidity may develop. 

A 100 c.c. graduated cylinder is now prepared for the unknown and a 
50 c.c. volumetric flask for the uric acid standard solution. Then 5 c.c. 
of uric acid standard (see page 545) (5 c.c. = 1 mgm. of uric acid) are meas- 
ured with a pipette into the 50 c.c. volumetric flask. To the uric acid 
standard 2 drops of a 5% solution of potassium cyanide, 2 c.c. of Folin- 
Macallum reagent (see page 120) and 20 c.c. of saturated sodium carbonate 
are added and, in 1 minute, water, to the 50 c.c. mark. To the precipitate 
in the centrifuge tube one now adds 2 drops of a 5% potassium cyanide 
solution (the tube is shaken so as to dissolve the precipitate), 2 c.c. of the 
Folin-Macallum reagent and then washes the contents of the centrifuge tube 
into a 100 c.c. graduate with from 15 to 20 c.c. of saturated sodium carbo- 
nate. More carbonate, i.e., 20 c.c, is used when the color is stronger than 
the standard and 15 c.c. when it is weaker, so that the unknown solution 
may be paler in colcr than the standard solution. From 40 to 60 seconds 
should now be allowed to pass before determining how much to dilute this. 

89 The 0.01N acetic acid is prepared by adding 0.6 c.c. of glacial acid to 1 liter of 
distilled water. This should be made up fresh at least each 2 weeks. 

90 For the preparation of alumina cream 500 c.c. of 8% aluminum acetate in acetic 
acid are precipitated with sodium bicarbonate (dry) until the solution is neutral to litmus 
paper. This is allowed to stand for 24 hours and the supernatant fluid decanted. This 
is repeated 6 times. On the sixth day the precipitate is filtered and stored in a jar with* 
the addition of 5 c.c. of chloroform. It is now ready for use. It should be kept in the 
ice-box for storage. 



THE BLOOD . 545 

It is then diluted with distilled water to 25, 50 or iooc.c, depending upon 
the depth of color obtained. Table II gives the data for working out the 
amount of uric acid present. 

Example. — Suppose the final dilution of the unknown was 25 and the 
reading was 42 . 42 in the table is equivalent to 1.24 mgms. This is divided 
by 4 because it is % as strong as the amount in the table (i. e., of 100) 
which equals 0.31 mgm. in 10 c.c. of blood (which is the amount of blood 
we started with). In 100 c.c. of blood we would have 10X 0.31 =3.1 mgms. 
of uric acid. 

Myers' Modification of Folin and Wu's Method. — To 5 c.c. of well-mixed 
oxalated blood in a 100 c.c. Erlenmeyer flask (or 100 c.c. glass stoppered cylinder) are 
added 35 c.c. of water (7 volumes), 5 c.c. of 10% sodium tungstate and finally 5 c.c. 
of exactly 0.75N sulphuric acid, rotating the flask all the time and then shaking thor- 
oughly. (There are some advantages in using 7 c.c. of blood since one secures rather 
quickly 40 c.c. of filtrate, the equivalent of 4 c.c. of blood, but satisfactory results may 
be obtained with but 2.5 or 3 c.c. of blood.) When the blood is properly coagulated the 
color of the coagulum will turn from pink to brown. If this change does not occur the 
coagulation is incomplete, due probably to too much oxalate, in which case 5% sulphuric 
acid may be added a drop at a time, shaking between each addition, until the coagulation 
is complete. Folin and Wu caution against an excess of sulphuric acid since this appar- 
ently precipitates some of the uric acid. If the mixture is now carefully poured upon 
the double portion of a filter just large enough to hold the mixture, the nitrate will 
probably come through perfectly clear; if not, the first portion may be returned to the 
filter. To prevent evaporation a watch glass may be placed over the top of the funnel. 
It is convenient to filter into a 50 c.c. graduated cylinder. 

When the filtration is complete the volume of the filtrate is recorded (ordinarily 
between 25 and 30 c.c.) and it is poured into a 50 c.c. centrifuge tube. Five cubic centi- 
meters of 5% silver lactate solution in 5% lactic acid are added, the fluid stirred and 
centrifuged. The supernatant fluid is then poured off. To the precipitate is added about 
3 c.c. of 10% sodium chloride in 0.1N hydrochloric acid (prepared by adding 1 c.c. of 
concentrated hydrochloric acid to 100 c.c. of 10% chloride solution). It is then stirred 
with a glass rod. One now adds 3 to 4 c.c. of water, 1 drop of 5% potassium cyanide, 
stirs again and centrifuges. This treatment sets the uric acid free from the precipitate. 
The supernatant fluid is then poured into an accurately graduated 25 c.c. cylinder. 

Into a similar 25 c.c. cylinder one introduces with an Ostwald-Folin pipette 1 c.c. 
of standard uric acid solution, 91 then 4 c.c. of the acidified sodium chloride solution and 
sufficient water to make the volume approximately the same as that of the unknown. 
One drop of 5% potassium cyanide is then added. 

To the standard is now added 1 c.c. of the uric acid reagent 92 and 0.5 c.c. to the 

91 Benedict's standard uric acid solution is prepared as follows: One dissolves 4.5 
gms. of pure crystalline hydrogen disodium phosphate and 0.5 gm. dihydrogen sodium 
phosphate in 200 to 300 c.c. of hot water. This is filtered and made up to about 250 c.c. 
with hot water. This warm, clear solution is poured on to 100 mgms. of uric acid sus- 
pended in a few cubic centimeters of water in a 500 c.c. volumetric flask. This is agitated 
until completely dissolved. Then at once exactly 0.7 c.c. of glacial acetic acid is added, 
the volume made up to 500 c.c, mixed and then 5 c.c. of chloroform added. One cubic 
centimeter of this solution contains 0.2 mgm. of uric acid. This solution should be freshly 
prepared every 2 months. 

92 Benedict's modification of the Folin-Denis uric acid reagent is prepared by boiling 
100 gms. of sodium tungstate, 20 c.c. of concentrated hydrochloric acid and 30 c.c. of 
85% phosphoric acid in 750 c.c. of distilled water for 2 hours, preferably under a reflux 
condenser, and then making the volume up to 1000 c.c. with water. 

35 



546 CLINICAL DIAGNOSIS 

unknown specimen. Then saturated sodium carbonate solution, 5 to 6 c.c, is added 
to the standard and 3 to 4 c.c. to the unknown. The color is allowed to develop for about 
5 minutes. The standard is then diluted to 20 to 25 c.c, and the unknown, if darker 
than the standard, to a similar depth of color. 

With this technic fairly deep blue colors are obtained even with normal blood, colors 
that can readily be matched in the colorimeter. The slight cloud that sometimes appears 
may easily be separated in the centrifuge. For the calculation the following formula 

may be used: — X ' — b =mgm. of uric acid in 100 c.c. of blood, in which 5 

R 25 B 

represents the depth of the standard (15 mm.), R the reading of the unknown, X the 
dilution of the unknown, 0.2 the strength of the standard in mgm. and B the number 
of cubic centimeters of blood to which the nitrate employed was equivalent. 

Estimation of Uric Acid with the Test Tube Colorimeter. — The technic 
described above readily lends itself also to use with the test tube colorimeter. 

Method. — Three cubic centimeters of well mixed oxalated blood are pipetted into 
a large test tube or small flask and 21 c.c. of water added. The precipitation of the pro- 
teins is now carried out according to the above described procedure of Folin and Wu, 
3 c.c. of the sodium tungstate and 3 c.c. of the 0.75 N H 2 S0 4 being used. 

In case it is necessary to add extra stilphuric acid this should be done 
carefully as an excess may lead to a precipitation of the uric acid. 

To 15 c.c. of the nitrate in a 20 c.c. centrifuge tube 93 are added 2 c.c. 
of 5% silver lactate in 5% lactic acid. After stirring, this is centrifuged. 
The supernatant fluid is poured off and discarded. To the precipitate is 
added about 1.5 c.c. of 10% NaCl in 0.1N HO and this stirred with a glass 
rod. Then 3 to 4 c.c. of water are added, the fluid stirred again and cen- 
trifuged. This treatment frees the uric acid from the precipitate. The 
clear supernatant fluid is poured into the right-hand tube of the colorimeter 
and one drop of 2.5% potassium cyanide added. 

Into the left hand tube of the instrument is introduced 1 c.c. of the 
standard uric acid solution (diluted 1 15 and therefore containing 0.04 mgm. 
of uric acid). Now 1.5 c.c. of the 10% NaCl in 0.1N HC1 is added and 
sufficient water to make the volume practically the same as that of the 
unknown. Then 1 drop of 2 . 5 % potassium cyanide is added. 

To both tubes now is added 0.3 c.c. of the uric acid reagent and 2 c.c. of saturated 
sodium carbonate solution. The color is allowed to develop for 3 to 5 minutes, the 
standard then diluted to 10 c.c. (20 c.c. if the unknown shows a weak color development) 
and the unknown to the same intensity of color as the standard, adding the water a 
drop or two at a time as one approaches the end point. 

To calculate the number of milligrams of uric acid per 100 c.c. of blood the following 
formula may be used, in which 5 represents the strength of the standard (0.2 or 0.4 mgm. 
to 100 c.c, depending on the dilution), D the dilution in cubic centimeters of unknown 
required to match the standard, and B the number of cubic centimeters of blood em- 
ployed. = mgm. uric acid per 100 c.c. of blood. If, for example, 15 c.c. of filtrate 
B 

are used, the equivalent of 1 .5 c.c. of blood, with a standard containing 0.4 mgm. diluted. 

93 Cylindrical trunnion cups may be substituted for the conical cups of the ordinary 
centrifuge. In these, test-tubes or special 20 c.c. centrifuge tubes may be employed. 
The cups work quite as well for the 15 c.c. conical centrifuge tubes. 



THE BLOOD 547 

to io c.c, and if the unknown was diluted to 17.1 c.c. to match the standard, the formula 

will work out as follows: 

0.4 X 17. 1 s 

— - - — = 4.6 mgms. to 100 c.c. 

1-5 

Creatinin in the Blood.— The creatinin of the blood, which is almost 
exclusively of endogenous formation, furnishes our best criterion as to the 
excretory power of the kidney (Myers) . The blood of normal persons con- 
tains from 1 to 2 mgms., as a rule about 1.2 mgms., per 100 c.c. of blood. 
While from 2 to 3 mgms. is abnormal yet we often find that much in 
the blood of patients who have no evidence of renal insufficiency. Above 
3 mgms. would mean a definite retention, over 4 mgms. would mean 
considerable impairment of renal function, while cases with 5 mgms. or 
more seldom show any improvement under treatment and practically all 
die in a rather limited time unless the renal condition be fairly acute, in 
which case recovery is possible. 94 

, The value in prognosis of creatinin determinations is well illustrated by 
those patients who with high creatinin content of the blood and with very 
few months to live yet are up and about, feel well and may show considerable 
clinical improvement. Some certainly show none of the subjective symp- 
toms usually associated with uremia. 

This was well illustrated by J. S., No. 8866, aged 33 years, admitted for " easy 
fatigability." He was pale, had albuminuric retinitis but practically no subcutaneous 
edema of any part of body, but for over a year had been rather slow of speech. His 
blood pressure varied from 180 to 210 mm. Hg., his spinal fluid gave a strong positive 
Wassermann reaction on all dilutions, his urine contained 2 gms. albumin per liter of 
urine and casts of all descriptions. His renal functional test, tried several times, gave no 
elimination of phenolsulphonephthalein in 2 hours and his blood creatinin, always high, 
was, when he left, 10.9 mgms. per 100 c.c. of urine. He died 10 days later. His doctor 
reported that he was quite clear mentally up to 1 hour of his death and had had no 
headache, no drowsiness and very little subcutaneous edema. 

For M. L. No. 9559, aged 45, see page 541. 

While preparing the specimen of blood for the determination of creatinin 
the standard solution should be prepared at the same time in order that 
the color may develop equally fast in both. Otherwise the results will 
be incorrect. 

The blood should be taken at least 14 hours after the last meal, that is, 
between 7 and 9 o'clock in the morning. Five cubic centimeters of blood 
measured in an Ostwald pipette are allowed to run to the bottom of a 50 c.c. 
centrifuge tube into which have previously been measured 20 c.c. of dis- 
tilled water. The pipette is washed by alternating drawing up and blowing- 
down this blood and water mixture, which is then stirred, to lake the cells. 
Dry picric acid (0.5 gm.) is added to precipitate the protein and the mix- 
ture thoroughly stirred. After standing for from 10 to 15 minutes, during 

94 For Myers' reports of cases with 5 mgms. or more, see Am. Jour. Med. Sci., 1919, 
clvii, 674, and the Jour, of Lab. and Clin. Med., June, 1920, vol. v, p. 566. 



548 



CLINICAL DIAGNOSIS 



which time it is stirred occasionally, the specimen is centrifugalized for 5 
minutes at about 1500 revolutions per minute and then filtered through a 
small filter paper into a clean, dry test tube. To 10 c.c. of this filtrate is 
added 0.5 c.c. of a 10% sodium hydroxide solution; while to 20 c.c. of a 
standard creatinin solution 95 is added 1 c.c. of 10% sodium hydroxide. 
Both are now allowed to stand for 10 minutes and then are read in 
the colorimeter. 

The formula for the computation of the result is as follows 189— read- 
ing X 0.0179 X 5 = the number of mgms. of creatinin per 100 c.c. of blood. 

Example. — Let us assume the reading in an experiment is 64. Then 89 
— 64 = 25X0.0179 = 0.4475X5 = 2. 2375 mgm?. 

Slightly less accurate results than these may be obtained by using as 
standard 0.25 N bichromate of potash solution (made by dissolving 12.28 
gms. of potassium bichromate in distilled water and making the solution 
up to 1 liter). When using this standard the nitrate is treated as in the 
preceding and the result is multiplied by 5. In this case Table V should 

b6USed - TABLE V* 



Estimation of Creatinin in the Blood with the Hellige Colorimeter 





Creatinin 




Creatinin 




Creatinin 


Colorimetric 


mgms. per 


Colorimetric 


mgms. per 


Colorimetric 


mgms. per 


reading 


dilution of 


reading 


dilution of 


reading 


dilution of 




100 c.c. 




100 c.c. 




100 c.c. 


40 


0.80 


57 


o-55 


74 


O.3I 


41 


O.78 


58 


o-54 


75 


O.3O 


42 


O.77 


59 


0.52 


76 


O.28 


43 


0-75 


60 


0.51 


77 


0.27 


44 


O.74 


61 


0.50 


78 


O.25 


45 


O.72 


62 


0.48 


79 


O.24 


46 


O.71 


63 


0.47 


80 


0.22 


47 


O.7O 


64 


o-45 


81 


0.21 


48 


O.68 


65 


0.44 


82 


0.20 


49 


O.67 


66 


0.42 


83 


0.18 


50 


O.65 


67 


0.41 


84 


0.17 


5i 


O.64 


68 


0.40 


85 


O.I5 


52 


0.62 


69 


0.38 


86 


O.I4 


53 


0.6l 


70 


0-37 


87 


0.12 


54 


0.60 


71 


o.35 


88 


O.II 


55 


O.58 


72 


o.34 


89 


0.10 


56 


o-57 


73 


0.32 


90 


O.O9 



* The table here given must be used when N/4 bichromate is used as a standard. Myers and Fine. 
Table copied from Gradwohl and Blaivas. 

Creatin of the Blood. — Under normal conditions the blood contains from 
3 to 7 mgms. of creatin per ioo c.c. While this is increased in the last stages 
of nephritis along with the other non-protein nitrogenous substances, the 
results of its determination have not as yet proven of value. (Myers.) 

For the determination of creatin plus creatinin 1 or 2 c.c. of the filtrate used for sugar 
or creatinin determinations is measured with an Ostwald-Folin pipette into a small 

95 The standard solution of creatinin is made by dissolving 15 mgms. of pure crea- 
tinin in 100 c.c. of a saturated solution of picric acid. 



THE BLOOD 549 

test-tube or a 10 c.c. graduate and autoclaved at 20 pounds pressure for 20 minutes. 
The solution is then cooled, made up to 8 c.c. with a saturated solution of picric acid 
and then 0.4 c.c. of a 10% sodium hydroxide solution added. The standard solution 
must be made up at the same time, as follows: To 20 c.c. of standard creatinin (made 
by dissolving 15 mgms. of pure creatinin in, and making the volume up to, 1 liter with 
saturated picric acid) add 1 c.c. of a 10% solution of sodium hydroxide (this should be 
added at the same time the 0.4 c.c. is added to the unknown). The unknown and the 
standard solutions are compared after standing for 10 minutes. The formula for com- 
putation of this result is as follows: 89— reading X 0.0 179X20= mgms. of creatinin 
plus creatin. 

Slightly less accurate results may be obtained by using 0.25 N potassium bichromate 
as a standard. 

If the accurate value of creatin is desired this is obtained by subtracting the value 
of creatinin previously determined from the creatin plus creatinin and multiplying the 
difference by 1.16. 

Example. — Let us assume that the reading in the specimen already examined for 
creatinin (see page 548) was 69. Then 89 — 69 = 20X0.0179=0.358X20 = 7.16 mgms. of 
creatinin + creatin = 7.16 — 2.2375 (e.g., mgms. creatinin) =4.9225X1.16 = 5.7101 mgms. 
creatin per 100 c.c. blood. 

Glucose in the Blood. — The blood of a normal person contains from 0.06 
to 0.11% (mean, 0.10%) of glucose. Strouse, 96 using a modified Kowarsky 
method, found that the normal blood sugar varied from 0.04 to 0.12%, 
average 0.084%. The nguies vary much in the same individual and depend 
on his diet. The curve has minima before meals and maxima 1 hour after 
those meals which contain carbohydrates. Above 0.12% is considered a 
hyperglycemia if the blood was drawn at least 14 hours after the last meal. 
After a meal it normally may rise to 0.17%. This increase begins at once 
(within half an hour) after the ingestion of glucose and a little later after 
a meal rich in starch. In 2 or 3 hours after such a meal the blood-sugar may 
fall below normal. 

In certain pathological conditions the percentage of the glucose of the 
blood is increased. Among these are diabetes mellitus, nephritis and hyper- 
throidism especially; some claim also, apoplexy, pneumonia, typhoid fever, 
tuberculosis, sometimes in cancer, after operations, after prolonged ether 
anesthesia and in arteriosclerosis. Attention should at this point be called 
to the importance of changes in the blood volume as explaining a rise or 
fall in the percentage of glucose. Epstein would limit the term hyper- 
glycemia to an increase in the total amount of glucose in the blood irre- 
spective of its percentage. 

Hypoglycemia would seem to be the result of subnormal endocrine 
functions since it has been noted in myxedema, cretinism, Addison's disease, 
pituitary disease and also in muscular dystrophy. 

In 12 cases of these conditions reported (Myers) the blood-sugar ranged from 
0.064 to 0.086%. 

96 Johns Hopkins Hosp. Bull., June, 1915, vol. xxvi, p. 211. 



550 CLINICAL DIAGNOSIS 

The question of a "renal threshold" above which glucose will pass over 
into the urine has attracted considerable attention. Various authors have 
placed this at. from 0.149 to 0.18% but this varies much and even in the 
same individual may be raised or lowered by various drugs and more es- 
pecially by conditions producing a diuresis which would tend to produce also 
a glycuresis. In diabetes mellitus, especially in the cases of long standing 
and particularly in those complicated by nephritis, there may be a hyper- 
glycemia of from 0.25 to 0.35% or more without glycosuria. This threshold 
is changed also by exercise. One of Joslin's patients showed that a liberal 
carbohydrate meal at noon would not produce a glycosuria if he exercised 
strenuously immediately thereafter. 

It is in diabetes mellitus that the amount of glucose in the blood has 
value in diagnosis, prognosis and treatment. In untreated cases of this 
disease the percentage usually lies between 0.20 and 0.40% but in well 
treated cases it may lie within normal limits. After a meal rich in carbo- 
hydrates the increase in the blood-sugar is slower than normal, reaching 
its maximum in 2 hours, while the decline occupies 8 or 10 hours. In cases 
before death with coma it may be above 0.40%, in cases with nephritis even 
0.80% and in 1 such case cf Joslin's 12 hours before death it was 1.37% 
(probably the highest figure on record). 

We take the liberty to draw the folic wing conclusions from Joslin's 
splendid work : First, the younger the patient the lower is the blood sugar 
apt to be (but the converse is not true). Second, the duration of the disease 
beais no necessary relation to the amount of sugar in the blood. (Therefore 
a case does not necessarily become more severe the longer it lasts.) Third, 
the presence of glucose in the urine is almost invariably preceded 24 hours 
before by an increase of the sugar in the blood. (On the other hand there 
may be a hyperglycemia of even 0.50% and no glycosuria.) Fourth, there 
is an intimate relation between blood sugar and the carbohydrate balance. 
This depends on the diet but also on the quantity of stored carbohydrate. 
Fifth, some very mild diabetics, especially those of the hereditary type, 
have early a low blood sugar per cent., even 0.11%, but a higher one later. 
Sixth, there is, in cases of long duration, a definite relation between the 
blood sugar and the patient's assimilation limit for glucose. The lower the 
tolerance for carbohydrates the higher the percentage of blood sugar and 
when the tolerance is distinctly high the blood sugar may be little above 
normal. There are, however, many exceptions to this rule. Seventh, in 
cases of long duration the blood sugar is seldom high no matter whether the 
patient's tolerance for carbohydrate is low or high. Eighth, those cases at 
first clinically severe but which become milder on treatment show corre- 
sponding changes in the blood sugar, but those cases which at first appear 
severe but later prove to be mild could not at first have been recognized as 
mild by blood analysis. Ninth, treatment is as a rule accompanied by a 
diminution in the blood sugar, although the patient's urine usually becomes 



THE BLOOD 551 

sugar- free before his blood sugar shows any particular diminution. This 
means that rigorous dietetic treatment should be continued for a long 
period after the urine is sugar-free. That is, the success of treatment de- 
pends on keeping the blood sugar withm normal limits as well as on keeping 
the patient's urine sugar- free. 

Renal diabetes on the other hand is a glycuresis due to a lowering of the 
renal threshold point below the level of the normal blood sugar. Unlike 
diabetes mellitus renal diabetes is the result of kidney disease. The renal 
signs are sometimes, but not always, those of nephritis. The condition 
would seem somewhat analogous to phlorizin glycosuria but resembles more 
uranium nephritis. Myers suggests that this condition may explain some 
cases of mild glycosuria which have not the classic symptom of diabetes 
mellitus. The blood sugar in Myers' cases ranged from .09 to .19% and 
the sugar in the urine from 0.3 to 1.3%. 

Determination of Carbohydrate Tolerance. — Killian's method 
(quoted from Myers) of determining the carbohydrate tolerance, which 
now promises to replace Naunyn's test for the assimilation limit (see page 
163) since the variable factor of the threshold point of renal excretion is 
eliminated, is as follows: The patient is given in the morning a standard 
breakfast consisting of 2 slices of bread, 1 egg in any form and 1 cup of 
water. Two hours later he empties his bladder and then drinks 200 c.c. of 
water. One hour later a specimen of blood and 1 of urine are taken as con- 
trols. The patient then ingests glucose in the form of a 50% solution, 
1.75 gms. (of glucose) per 1 kg. of body weight. The blood sugar is then de- 
termined each hour for 3 or 4 hours and its curve plotted. The urine for 
the 24 hours following the meal is collected and the presence of glucose de- 
termined. In one modification of the Goetch test for hyperthyroidism the 
determination of blood glucose is made. The patient is given 100 gms. of 
glucose by mouth and then 8 minims of adrenalin are injected under the 
skin and the blood sugar followed each hour. In a positive test there will 
be a definite rise of the blood sugar and sometimes a glycosuria. 

Case No. 10155, a girl 17 years of age with hyperthyroidism, showed during the 
Goetch test a rise in systolic blood pressure from 1 15 to 142 mm. Hg. in about 1 1 minutes. 
It returned to 1 18 in approximately 1 hour. The pulse rose from 100 to 120 per minute. 
The subjective symptoms were very marked; tremor, which was marked and general- 
ized, and a numb feeling which lasted about 50 minutes. The blood sugar on the fasting 
stomach was 0.12%. One hour after the ingestion of 100 gms. of glucose by mouth 
this was 0.19% and at the end of about 5 hours, 0.18%. Hourly urine specimens were 
sugar-free. The blood creatinin was 2.27 mgms. per 100 c.c. and basalmetabolism 45.7 
cal. per square M. per hour. 

In case No. 9998 the Goetch test showed a rise in systolic pressure from 130 to 138 
mm. Hg. where it remained for 2 hours. The pulse-rate increased from no to 128 per 
minute. The blood sugar rose from 0.17% to 0.20% after 100 gms. of glucose by mouth. 
In 234 hours it was 0.18% and in 4^ hours it had fallen to 0.13%. All urine 
voidings were negative for sugar. This case had a negative Goetch cutaneous reaction. 



552 CLINICAL DIAGNOSIS 

All the clinical signs of hyperthyroidism were present. The basalmetabolism test gave 
48.4 cal. per kgm. per 24 hours, or an increase of 87%. 

Case No. 9782, of adenoma of the thyroid with toxic symptoms, gave a positive 
Goetch test. The blood-pressure rose from 112 to 128 mm. Hg. in about 35 minutes 
and returned to normal in 1 hour. The pulse rose from 102 to 120 per minute. There 
were no subjective symptoms. The Goetch cutaneous was negative. But the blood 
sugar, which before the test was 0.16%, rose in 1 hour after the ingestion of 100 gms. of 
glucose by mouth to 0.21%. The 2d hour it dropped to 0.17%, the 4th hour it read 
practically the same and at the 6th hour it read 0.16%. The urine passed the second 
hour, when the blood sugar was the highest, was negative for sugar but glucose was 
present in the next 2 hourly specimens. The later specimens were negative. The blood 
creatinin was 1.7 mgms. per 100 c.c. 

Quantitative Determination of Glucose in the Blood. — (Lewis-Benedict 
method modified by Gradwohl and Blaivas) . The blood should be taken in 
the post absorptive stage, i.e., 14 hours after the last meal, which usually is 
between 7 and 9 in the morning. The estimation of the glucose of the blood 
should be begun as soon as possible after the blood is drawn, otherwise 
the specimen will deteriorate rapidly. One measures 3 c.c. of the filtrate 
(see page 547) into a sugar tube, a graduated test tube on which are 
marked 1, 4, 10, 15, and 20 ccs., adds 1 c.c. of saturated sodium carbonate 
(prepared by dissolving 220 gms. of anhydrous sodium carbonate in 1000 c.c. 
of distilled water) and mixes it well. The test tube containing this mixture 
is then immersed in a large beaker of water which is then boiled over a free 
flame for about 1 5 minutes and then allowed to cool. This cooled solu- 
tion is then so diluted with distilled water that it will be weaker in color 
than the standard picramic acid solution with which it is to be compared 
in the colorimeter. To this end it is diluted to 10, 15, or 20 c.c. (see marks 
upon the graduated sugar tube. In average normal cases a dilution to 
10 c.c. will suffice but in cases of hyperglycemia it is often necessary to 
dilute to 1 5 c.c. or even to 20 c.c. It is now compared in the colorimeter with 
a wedge of standard picramic acid. The readings should be made as 
rapidly as possible since the color will soon change. 

The standard picramic acid solution is a staple solution and is prepared 
as follows: Dissolve 0.1 gm. of picramic acid and 0.2 gm. anhydrous 
sodium carbonate in 30 c.c. warm distilled water and dilute to 1 liter. 

Example. — If the dilution was to 10, multiply the difference between 
the reading and 100 by 0.002 ; if to 15, by 0.003 \ if "to 20, by 0.004; while if 
the dilution is to 25, multiply by 0.005 I etc. 

Identical results maybe obtained by using the data presented in Table VI, 
providing the estimation was made on the basis of a dilution of 10. If 
it was diluted to 15 c.c, multiply the result by 1.5; to 20 c.c, multiply 
by 2 ; etc. 

Creatinin will be estimated by this method as glucose but the error is 
not great since creatinin very seldom exceeds 20 mg. per 100 c.c of blood. 
To lessen this error Myers proposes the following method in which a lower 
dilution of blood is used. 



THE BLOOD 

TABLE VI* 



553 



Estimation of Blood Sugar with Hellige Colorimeter 



Colorimetric 


Blood sugar in 


Colorimetric 


Blood sugar in 


Colorimetric 


Blood sugar in 


reading 


per cent 


reading 


per cent 


reading 


per cent. 


25 


O.150 


45 


O.IIO 


65 


O.070 


26 


O.I48 


46 


O.I08 


66 


O.068 


27 


O.I46 


47 


O.I06 


67 


O.066 


28 


O.I44 


48 


O.IO4 


68 


O.064 


29 


O.I42 


49 


O.I02 


69 


O.062 


30 


O.I4O 


50 


O.IOO 


70 


O.060 


31 


O.138 


5i 


0.098 


71 


O.058 


32 


O.I36 


52 


0.096 


72 


O.056 


33 


O.I34 


53 


0.094 


73 


O.054 


34 


O.I32 


54 


0.092 


74 


O.052 


35 


O.130 


55 


0.090 


75 


O.O5O 


36 


0.128 


56 


0.088 


76 


O.O48 


37 


O.I26 


57 


0.086 


77 


O.O46 


38 


O.I24 


58 


0.084 


78 


O.O44 


39 


O.I22 


59 


0.082 


79 


O.O42 


40 


O.I20 


60 


0.080 


80 


O.O4O 


41 


0.II8 


61 


0.078 


81 


O.O38 


42 


O.Il6 


62 


0.076 


82 


O.036 


43 


O.II4 


63 


0.074 


83 


O.O34 


44 


O.II2 


64 


0.072 


84 


O.032 



* Myers and Fine. Table copied from Gradwohl and Blaivas. 

Myers and Bailey's Method.— To 8 c.c. of distilled water in a 20 c.c, cylindrical 
centrifuge tube are added 2 c.c. of well mixed oxalated blood (or oxalated plasma). 
This is then stirred with a glass rod until the blood is well hemolyzed after which about 
0.5 gm. of dry picric acid (sufficient to precipitate completely the proteins and to make 
a saturated solution) is added. 

The mixture is thoroughly stirred at intervals of several minutes until it is uniformly 
yellow, it is then centrifugalized and the supernatant fluid filtered into a dry test tube 
through a small 4 cm. filter paper. 

Three cubic centimeters of the filtrate are measured with a pipette into a tall narrow 
sugar tube (12X200 mm.) graduated to 3, 4, 10, 15, and 20 c.c. Then 1 c.c. of saturated 
(22%) sodium carbonate is added and the tube heated in a beaker of boiling water for 
15 or 20 minutes. Simultaneously 3 c.c. of a 0.020% solution of glucose in saturated 
picric acid (this keeps permanently) is treated with a similar amount of sodium carbonate 
in a sugar tube and heated for the same time as and with the unknown. This serves as 
the standard. The yellow sodium picrate is converted by the heat and alkali to reddish 
brown sodium picramate in proportion to the amount of sugar present. The solutions 
are now cooled to room temperature either by allowing the tubes to stand or by placing 
them in a beaker of water. The solution in the standard tube is made up to exactly 
10 c.c. with water and the contents of the other tube then diluted to some definite volume 
(as 10, 15 or 20 c.c.) the color of which approximates the color of the standard. It is 
best to allow a little time to elapse before color comparisons are made. These fluids 
are then compared in a colorimeter. The standard may conveniently be set at the 
15 mm. mark. 

Since the 3 c.c. of filtrate is the equivalent of 0.6 c.c. of blood and the 3 c.c. of stand- 
ard solution contains 0.6 mgm. of glucose, the proportions are the same as if 100 c.c. 
of blood and 0.1 gm. of glucose had been employed. 



554 CLINICAL DIAGNOSIS 



SXD Xo.i . , U1 , 

— =per cent, of blood sugar. 

J\ X io 



5 = depth of standard (15 mm.). 

D = dilution of unknown. 

0.1 = strength of standard in grams calculated on the basis of 100 c.c. of blood. 

R= reading of unknown. 

10 = dilution of standard. 

To test the purity of the picric acid Folin and Doisy recommend the following 
method. 97 To 20 c.c.of a saturated solution of picric acid add 1 c.c. of 10% NaOH 
and allow it to stand for 15 minutes. The color of the alkaline picric solution thus ob- 
tained should not be more than twice as deep (measured in a colorimeter) as the color 
of the saturated acid solution. 

For the determination of acetone, diacetic acid and hydroxybutyric acid 
in the blood see page 197. 

Diastatic Activity of the Blood. — The diastatic activity of the blood is 
increased in conditions of hyperglycemia and therefore in diabetes and 
nephritis, with the exception of the cases of diabetes due to lues. The 
increase of diastase in the blood of nephritics is probably due to a de- 
creased elimination of this in the urine. The figures for diastase in normal 
individuals is given by Myers 98 as 16 and 17%. In diabetes it may range 
from 24 to 74% running in general parallel to the blood sugar. Myers and 
Killian " suggested that the increase of blood diastase may be an impor- 
tant factor in the production of a hyperglycemia. 

Determination of the Diastatic Activity of the Blood. — Two 2 ex. samples 
of oxalated blood, 1 for control, are measured into two 20 c.c. cylindrical centrifuge tubes. 
The contents of the control is made up to 10 c.c. with distilled water and the other tube 
to 9 c.c. Both tubes are now placed in a water bath the temperature of which is main- 
tained constantly at 40 C. As soon as the contents of the tubes has been brought to 
this temperature 1 c.c. of 1 % soluble starch (which does not contain over 6% of reducing 
sugar; this reagent is tested before it is used) is added to the second tube, the contents 
mixed and the tubes incubated at 40 C. for exactly 15 minutes. At the end of this 
time about 0.5 gm. of dry picric acid is at once added to each tube and the mixtures 
stirred. When the proteids have been precipitated the tubes are centrifugalized and the 
yellow supernatant fluids filtered. The sugar in 3 c.c. portions of each of the filtrates 
is now estimated by the method described on page 552. Correction is made for the sugar 
originally present in the blood (with the aid of the control) and for the reducing bodies 
of the soluble starch if any existed. 

The results are recorded in terms of the percentage of the soluble starch (10 mgms.) 
transformed to reducing sugars (calculated as glucose) by the 2 c.c. of blood. 

— — — — = mgms. of reducing sugar in terms of glucose for 2 c.c. of blood. 

5 = depth of standard (15 mm.). D= dilution of unknown in cubic centimeters. 

2.0 = strength of standard in milligrams compared to 2 c.c. of blood. R= reading of 
unknown in millimeters. 10 = dilution of the standard. 

The difference between the results of the control and the test specimens multiplied 

97 Jour. Biol. Chem., 1916-17, xxviii, p. 349. 

98 The Jour, of Lab. and Clin. Med., July, 1920, vol. v, No. 10, p. 640. 

99 Jour. Biol. Chem., 1917, vol. xxix, p. 179. 



THE BLOOD 



555 



by io (10 mgms. of the soluble starch were used) gives the percentage transformation 
of the starch to reducing sugar provided the soluble starch requires no correction. For 
example, a diabetic blood with 0.26% blood sugar would give a control of 5.2 mgms. 
(since 2 c.c. were used). Suppose that the specimen gave 9.8 mgms. reducing sugar 
and that the soluble starch contained 6% reducing substance, 9.8 — (5.2 +0.6) X 10 = 40, 
the diastatic activity. 

Blood Lipoids. 10 ° — Joslin, quoting Bloor, classifies the lipoids in the 
blood as: (1) Glycerides of the fatty acids, especially of oleic, stearic and 
palmitic acids. (2) Lecithin-like bodies, which are compounds of glycerin, 
fatty acid, phosphoric acid and cholin and called often ''phosphatides." 
(3) Cholesterol, a stable secondary alcohol belonging to the terpene series 
and containing one double bond. (4) Cholesterol esters, which are combi- 
nations of cholesterol and a fatty acid. 

The lipoids may be classified also by grouping the constituents of the 
above bodies, as: (1) Total fatty acids, which vary in the whole blood 
between 0.29 and 0.42%; (2) lecithin, which varies from 0.28 to 0.33%; 
and (3) cholesterol which varies from 0.19 to 0.25%. Cholesterol in all 
cases runs parallel to the fatty acids, therefore its determination should 
give valuable information regarding the lipoid content of the blood, but not 
as valuable as that of the total fatty acids, since the lowest figures of the 
latter in the diabetic blood were at the upper limits of normal, while those 
for cholesterol overlapped the normal range. 

We copy from Joslin the table (for which he gives Bloor credit) of li- 
poids of normal blood. 

Lipoids of Normal Blood * 





Total fatty acids, 
grams per 100 c.c. 


Lecithin, 
grams per 100 c.c. 


Cholesterol, 
grams per 100 c.c. 




Whole 
blood 


Plasma 


Cor- 
puscles 


Whole 
blood 


Plasma 


Cor- 
puscles 


Whole 
blood 


Plasma 


Cor- 
puscles 


Per cent, variation of 
high above average.. . 

Highest normal 

Average (19) t normals. 

Lowest normal 

Per cent, variation of 
low below average . . . 


14.O 
O.42 

0.37 
O.29 

22.0 


20.0 

O.47 

0-39 
O.3O 

23.O 


32.0 
0-45 
0.34 

O.27 

21.0 


IO.O 

0.33 

O.3O 
O.28 

7.0 


24.O 
0.26 
0.21 
0.17 

I9.O 


14.O 
O.48 
O.42 

o-35 
17.0 


I2.0 
O.25 
0.22 
O.I9 

I4.O 


35-0 
0.31 
0.23 
0.19 

17.0 


20.0 
O.24 
0.20 
O.17 

15.0 



*The results of the analyses of blood lipoids of both males and females have been combined in 
this table. 

t The number of analyses are given in parentheses. 

The influence of food on the blood fat is striking. Fasting and narcosis 
cause an increase in the blood lipoids (probably only if the animal was pre- 
viously well fed) . 

The total fatty acids of the blood have been found increased in nephritis, 
pneumonia, pregnancy and in the experimental anemias of animals. 

100 Joslin, Treatment of Diabetes Mellitus, Second Edition, 19 17, page 96. 



556 



CLINICAL DIAGNOSIS 



Lecithin appears to be an intermediate stage in the metabolism of 
fat, formed in the red corpuscles from the fat absorbed from the plasma. 
Lecithin has been found increased in nephritis and in leukemia (in the 
corpuscles). 

Cholesterol (cholesterin) does not seem to be increased by the mere in- 
gestion of fat, but is increased in narcosis, alcoholism, pregnancy, jaundice 
and nephritis and is decreased in cachexia. Schmidt 101 found in normal 
persons the following figures: 

Cholesterin in Normal Blood-Sera 



Ag. 



Observa- 
tions 



Amount in grams per liter 



IO-I9 
20-29 

30-39 
40-49 

50-59 
60-69 



I.40 

1. 60-1. 27-1. 75-1. 45-1 .50 

1. 65-1 .25-1 .35 

1.20 

1 -45-i -55-i -20 

1. 30-1. 20-1. 30-1. 35 



For a series of very careful and interesting studies on cholesterol the 
reader is referred to those of Luden. 102 

By lipemia is meant an increase of the visible fat of the blood plasma, 
yet the condition is more one of lipoidemia since it is due to an increase of 
the glycerides of the fatty acids, of lecithin and cholesterol. A lipemia is 
physiological in sucklings, in very obese persons, in some pregnant women 
and in adults after a heavy meal. 

Recently, however, the term has been reserved for those cases in which 
the plasma is milky 14 hours after the last meal. Severe diabetes mellitus 
is the only disease in which lipemia is frequent enough to be of special 
significance (see page 452). Futcher 103 has contributed an article on this 
subject. Fraser 104 reported a case with 16.44% of fat in the blood and 
18.94% in the pleural exudate. The record case is Fischer's, with 18.129% 
in the blood . 

In diabetes mellitus the blood lipoids, chiefly of the plasma, were in- 
creased 50% in 26 of 28 cases. A true lipemia, or milky plasma, due to 
the presence of an abundance of fatty globules which in extreme cases may 
even rise as a cream, may easily be demonstrated soon after a rich meal in 
a severe case, but is rare in cases under treatment if the blood is drawn 14 
hours after the last meal. 

The amounts obtained by chemical analysis are shown in the following 
table copied from Joslin. 



101 Arch, of Int. Med., Jan., 1914, vol. xiii, p. 121. 

102 Jour. Lab. and Clin. Med., 1916, vol. i, p. 662; 1917, vol. iii, p. 93. 

103 Jour, of Am. Med. Assoc., Oct. 21, 1899. 

104 Scot. Med. and Surg. Jour., 1903, p. 200. 



THE BLOOD 557 

Comparison of Blood Lipoids of Normal and Twenty-eight Diabetic Individuals* 





Total fatty acids, 
gms. in ioo c.c. 


Lecithin, 
gms. in 100 c.c. 


Cholesterol, 
gms. in 100 c.c. 




Whole 

blood 


Plasma 


Cor- 
puscles 


Whole 
blood 


Plasma 


Cor- 
puscles 


Whole 
blood 


Plasma 


Cor- 
puscles 


Diabetic extremes . . 
Diabetic average ( 31 ) 

Normal average ( 19 ) 
Normal extremes . . . 


.41-76 
■52 

■37 
.29-42 


.46-. 93 

■59 

■ 39 
• 30-47 


.33-62 

■43 

■ 34 
• 21-. 45 


.26-50 
.36 

• 30 
■28-. 33 


.17-.48 
■30 

.21 
.17-.26 


.32-. 60 

.46 

•42 

.35--48 


.19-. 44 
■ 29 

.22 
.19-25 


.16-.65 

■ 36 

■ 23 
.19-31 


.17-24 
.20 

.20 
.17-24 



*Compiled by Joslin from tables of W. R. Bloor, Jour. Biol. Chem., 1916, xxvi, p.424 

Joslin concludes that the more severe the diabetic condition the more 
abnormal the quantities of lipoids in the blood. While in general the re- 
lations between the various lipoids in diabetes are practically the same as in 
normal individuals yet there is a tendency for the total fatty acids to in- 
crease out of proportion to the other constituents. The best explanation 
for this increase in the blood lipoids is a disturbance of the fat-burning 
mechanism and the consequent accumulation of metabolites. 

The presence of visible fat in lipemia, seen so often in the patients on the 
diets formerly so popular but so seldom in patients under Allen's treatment, 
would seem to depend directly on the abundant fat of the diet which con- 
tained, as little as possible of carbohydrates. 

The fat will remain in solution until the lecithin-forming function fails. 

The lipemia sometimes present in blood in wmich there is not a suf- 
ficient increase of the blood lipoids to warrant its presence is explained by 
changes which take place in a clear diabetic plasma after it has stood more 
than a few hours. 

Joslin was unable from a study of his cases to demonstrate any definite 
relation between blood lipoids and acidosis, the duration of the disease, 
the prognosis or the amount of blood sugar. 

Quantitative Determination of the Lipoids of the Blood. — 
(Joslin, p. 207.) The blood should be obtained 14 hours (from 8 to 16) 
after the last meal and the analysis begun as soon as possible (not later 
than 2 hours) after the blood is drawn. 

Three cubic centimeters of freshly drawn and well mixed blood are 
run in a slow stream of drops into a graduated flask containing about 
80 c.c. of a mixture of 3 parts alcohol and 1 part ether (both redistilled), 
which is kept in constant motion by rotating the flask. The solution is 
raised to boiling by immersing the flask in a water bath (with frequent 
shaking to prevent superheating), then cooled to room temperature, 
made up to volume with the alcohol-ether mixture, mixed and filtered. 
This extract in tightly stoppered bottles in the dark will keep unchanged 
for several months. 

From 5 to 20 c.c. (ordinarily 10 c.c.) of the extract, i.e., that amount 
which would contain about 2 mgms. of "fat," are measured with a pipette 



558 CLINICAL DIAGNOSIS 

into a small beaker and saponified by evaporation just to dryness with 2 
c.c. of iN sodium ethylate (made by dissolving cleanedjmetallic sodium 
in absolute alcohol). After evaporation is complete 5 c.c. of alcohol-ether 
are added and the mixture heated slowly to boiling. 

A similar solution is prepared by measuring 5 c.c. of the standard fat 
solution (see below) into a beaker and heating to boiling as above. Fifty 
cubic centimeters of distilled water are now added to each beaker and the 
solutions mixed by stirring, taking care that all the material in the saponi- 
fication beaker is dissolved. To standard and test solutions are added, as 
nearly simultaneously as possible, 10 c.c. of dilute (1 to 4) hydrochloric 
acid and the solutions allowed to stand 5 minutes after which they are 
transferred to the comparison tubes of the nephelometer. If bubbles appear 
on the walls of the tubes they should be removed by inverting the tubes 
2 or 3 times. The movable jacket of the standard tube is set at a convenient 
point, generally 50 mm. (Richard's nephelometer), and comparisons made by 
adjusting the jacket on the test solution until the images of the 2 solutions 
show equal illumination. Not less than 5 readings are made, alternately 
from above and below, and the average taken as the correct reading. 

The standard solution used is an alcohol-ether solution of pure triolein 
of which 5 c.c. contain about 2 mgms.of fat. Freshly redistilled absolute 
alcohol and pure dry ether should be used for making the standard solution. 

Cholesterol. — For the determination of the cholesterol of the blood 
2 c.c. of the whole blood, plasma, or serum are run in a slow stream of drops 
from a pipette into a 100 c.c. graduated flask containing about 75 c.c. of a 
mixture of redistilled alcohol, 3 parts, and ether, 1 part. The contents of 
the flask should be kept in motion during the process so that there can be 
no clumping of the precipitated material. The contents of the flask is now 
heated to boiling by immersion in a water-bath (with constant shaking to 
avoid super-heating) , cooled to room temperature, filled to the 100 c.c. mark 
with the alcohol-ether mixture, mixed and filtered. This filtered liquid will 
keep tightly in a stoppered bottle in the dark unchanged for a consider- 
able time in case it is not convenient to complete the determination at once. 

The slow addition of blood to a large quantity of alcohol-ether will 
precipitate the protein material in finely divided form and so brief heating 
is adequate for the complete extraction of serum or plasma. The extraction 
is not so complete in the case of whole blood and yet whole blood is to be 
preferred because of the higher values obtained. 

One now measures 10 c.c. of the alcohol-ether extract into a small flat- 
bottomed beaker and evaporates just to dryness over a water-bath or 
electric stove. Overheating would produce a brownish color which would 
pass into the chloroform and render the subsequent determination dif- 
ficult or impossible. The cholesterol is extracted from the dry residue by 
boiling it out 3 or 4 times with successive small portions, 3 c.c, of chloro- 
form each of which is allowed to boil down to half its volume or less and then 



THE BLOOD 



559 



decanting into a 10 c.c. glass-stoppered graduated cylinder. The combined 
extracts after cooling (5 c.c. or less) are then made up to 5 c.c. The solution 
should be colorless but not necessarily clear, since a slight turbidity will 
disappear on adding the reagents. 

To this solution are added 2 c.c. of acetic anhydride and 0.1 c.c. of con- 
centrated sulphuric acid, the fluids mixed and placed in the dark for 10 
minutes to allow for the development of the color. The specimen is then 
compared in the colorimeter (Hellige) with a standard cholesterol solution 
which one has been making while preparing the specimen in order that 
the colors of the unknown and the standard solutions may develop at the 
same time since the colors fade rather rapidly. It is very important that 
the wedge and the cup of the colorimeter be perfectly dry. 

For the preparation of the standard cholesterol solution 2 c.c. of an 0.08% 
freshly prepared chloroform solution of cholesterol is pipetted into a dry, 
accurately graduated 25 c.c. cylinder and made up to 10 c.c. with chloro- 
form. One then adds 4 c.c. of acetic anhydride and 0.2 c.c. of concentrated 
sulphuric acid. 

An aqueous solution of Naphthol Green B can also be used as a standard. 
The cholesterol content of 0.2 c.c. of blood, serum, or plasma, can be ob- 
tained from Table VII. This table is suitable for both standards (pure 
cholesterol or Naphthol Green B). The result multiplied by 500 will give 
the percentage of cholesterol. 

TABLE VII* 
Estimation of Cholesterol with the Hellige Colorimeter 





Cholesterol 




Cholesterol 




Cholesterol 


Colorimetric 


mgms. dilution 


Colorimetric 


mgms. dilution 


Colorimetric 


mgms. dilution 


reading 


of 5 c.c. 


reading 


of s c.c. 


reading 


of 5 c.c. 


15 


O.74 


35 


0.57 


55 


0.40 


16 


0-73 


36 


O.56 


56 


O.4O 


17 


O.72 


37 


0.55 


57 


0-39 


18 


O.71 


38 


0-55 


58 


0.38 


19 


O.70 


39 


0-54 


59 


o.37 


20 


O.69 


40 


0.53 


60 


0.36 


21 


O.69 


4i 


O.52 


61 


0-35 


22 


O.68 


42 


O.51 


62 


o.35 


23 


O.67 


43 


O.50 


63 


o.34 


24 


O.66 


44 


O.50 


64 


o.33 


25 


O.65 


45 


O.49 


65 


0.32 


26 


O.65 


46 


O.48 


66 


0.31 


27 


O.64 


47 


O.47 


67 


0.30 


28 


O.63 


48 


O.46 


68 


0.30 


29 


O.62 


49 


0-45 


69 


0.29 


30 


O.61 


50 


0-45 


70 


0.28 


31 


O.60 


5i 


O.44 


71 


0.27 


32 


0-59 


52 


0-43 


72 


0.26 


33 


o-59 


53 


O.42 


73 


0.25 


34 


0.58 


54 


O.41 


74 


0.24 



* Myers and Fine, Table copied from Gradwohl and Blaivas. 






560 CLINICAL DIAGNOSIS 

Suppose the reading is 60. This would equal 0.36 mgm. of cholesterol in 
0.2 c.c. of blood, plasma, or serum. Then -0.3 6*X 500= 180 mgms. or 0.18%. 

For the preparation of Naphthol Green B, dilute 2 c.c. of a 0.1% aqueous 
solution of the dye to 17 c.c. with distilled water. The diluted solution 
appears to keep for a little time, while the concentrated solution apparently 
will keep for a considerable time. The permanency of the solution and the 
fact that the color is practically identical with that obtained from choles- 
terol makes the standard very convenient. Myers and Fine advise, how- 
ever, to restandardize each new solution. 

The method which Schmidt (q.v.) used, which was modified from 
Grigaut, and Weston and Kent, is as follows: 

Two cubic centimeters of blood-serum were placed in a pressure bottle of 150 c.c. 
capacity. To this was added 20 c.c. of 1% solution of potassium hydroxide dissolved 
in 50% alcohol. The cover was screwed down and the whole placed in a boiling water- 
bath for from 15 to 20 minutes. After removal from the water-bath, the contents of the 
pressure bottle were allowed to cool and were then shaken with 50 c.c. of ether. The 
ether was decanted into a separating funnel and 30 c.c. more of ether was added to the 
remaining fluid and shaken vigorously as before. This was again decanted into the 
funnel and any aqueous liquid which separated from the ether removed. The ether 
was then washed with 80 c.c. of distilled water. The ethereal extract, after separation 
from the wash water, was poured into an evaporating dish and evaporated nearly to 
dryness, leaving behind the small yellow oily droplets containing the cholesterin. Five 
cubic centimeters of chloroform (Merck's blue label) was then placed on the evaporating 
dish and rotated carefully so as to dissolve the oily droplets as completely as possible. 
This chloroform solution was transferred to a convenient receptacle, and a second 5 c.c. 
of chloroform was then used to wash the evaporating dish and were added to the first 5 c.c. 

The amount of chclesterin in the chloroform solution was determined 
by comparing its color reaction with those of cholesterin chloroform solutions 
of known strength. These standard solutions were prepared by dissolving 
Merck's cholesterin in pure chloroform (Merck's blue label) and by prepar- 
ing a series of dilutions of varying strength so that the amount of cholesterin 
in 1 c.c. of chloroform varied from 0.0002 to 0.0004 gm. A set of test-tubes 
of uniform bore were arranged in a rack and 1 c.c. each of these known 
solutions of cholesterin were added in ascending series, as was also 1 c.c. 
of the cholesterin solution prepared from the blood-serum. It was often 
necessary to dilute the latter so that the amount would fall within the scale 
of the known quantities. To each tube was now added 0.1 c.c. of concen- 
trated sulphuric acid, and the mixture was thoroughly shaken. After 30 
minutes 1 c.c. of chloroform was added to each tube and after being placed 
for 1 5 minutes in a dark room, the color of the unknown solution was com- 
pared with that of the known. This method was found to be exceedingly 
sensitive and differences in the original fluid of 0.00005 g 111 - P er c - c - could be 
detected. Successive readings from the same serum also gave concordant 
results, and even after several days' standing little or no diminution in the 
amount of the cholesterin in the serum could be discovered. 



THE BLOOD 561 

Quantitative Determination of Fat in the Blood. — This 
method is valuable when large amounts of blood can be obtained. The 
method chosen by Bonninger, in Salkowski's laboratory 105 is as follows: 
From 5 to 30 grn-s. of blood are mixed with 10 to 20 volumes of 96% alcohol, 
the precipitate ground fine and then allowed to stand one or two days. It 
is then filtered, the precipitate extracted several times with alcohol in the 
same way, then twice with from 5 to 10 volumes of ether, digesting it each 
time for 1 day. All these extracts are then combined, evaporated, the 
residue repeatedly taken up in absolute alcohol, and this evaporated, then 
filtered, dried, and weighed. Extracting twice with alcohol alone would 
give 96% of the total fat. 

BACTERIOLOGY OF THE BLOOD 

Assuming on the part of the worker a thorough training in bacterio- 
logical technic, we shall consider below only such special points as may be 
useful in the study of the blood. 

The success of blood-cultures is in part dependent upon the obtaining 
of a sufficient quantity of blood for observation, 1 5 or 20 c.c. being the usual 
amount withdrawn, and on its quick dilution in a large volume of physio- 
logical salt solution or other dilute medium in order to protect the organisms 
from the bactericidal properties of the blood which increase after it is drawn. 
In general, it is obtained from the median basilic or cephalic vein although 
a smaller vein on the dorsum of the hand or foot may be used. Incision of 
the skin to expose the vein is not necessary unless the patient is very fat 
01 edematous. For typhoid cultures in bile mediums where only a small 
amount of blood is needed the finger-tip or the lobe of the ear may be 
cleansed with soap, then alcohol and ether, then coated with collodion. 
The skin is then punctured through the collodion and the blood allowed 
to drip directly into the tube. 

If the skin is carefully cleansed the chance of contamination by skin 
organisms is negligible. Those very careful scrub the site of operation 
(puncture) with green soap and hot water, then rub it over with Harrington's 
solution, wash it with ether and alcohol and then cover it with a wet 
bichloride (1 in 1000) compress. But more merely paint the skin a deep 
brown color with the tincture of iodine and then may or may not rub it 
clean with 95% alcohol. This is quite as satisfactory as the more elaborate 
method. 

If a syringe is to be used it snould have a capacity of at least 20 c.c. and 
a glass barrel which is perfectly true. Instead of the ordinary washer for 
the needle a piece of soft black rubber tubing may be cut and, after perfo- 
rating it with a pin, slipped over the nipple. This withstands boiling longer 
and gives a tighter joint. A fresh rubber should be used for each culture. 
The steel, or better irido -platinum, needle should be short and stiff, sharp 

105 Zeits. f. klin. Med., 1901, vol. xlii. 
36 



562 CLINICAL DIAGNOSIS 

and of moderately large caliber. The syringe, needle and a pair of forceps 
are sterilized in the autoclave or by boiling them for 15 minutes. The 
forceps are used in putting the needle on the syringe. The more popular 
apparatus now is that introduced by Rosenau (Fig. 129) which consists of a 
milk bottle of 200 c.c. capacity almost filled with the liquid medium and 
closed with a doubly perforated stopper, the one for the tube carrying the 
needle, the other for a tube through which suction can be made. By this 
means contaminations are avoided and the blood will at once mix with the 
medium. A moderately tight bandage is placed on the arm proximal to the 
site of operation to distend the vein. The skin may be anesthetized with 
ethyl chloride spray, but this is seldom necessary. The needle is plunged 
through the skin directly into the vein and the piston is drawn slowly, al- 
lowing the syringe to fill with blood, or suction made until the required 
amount of blood is obtained. The bandage should be removed before the 
needle is withdrawn since this will prevent bleeding. After withdrawal, 
the needle and washer of the syringe are removed and the media inoculated 
quickly. The tip of the syringe should be passed through the flame of an 
alcohol lamp before inoculating each tube. 

If the blood must be sent to a laboratory in order that cultures may be 
made, it may be discharged from the syringe into tubes or flasks contain- 
ing an equal volume of sterile isotonic ammonium oxalate (ammonium 
oxalate 2, sodium chloride 6 and water 1000) solution. 

Agar tubes melted and cooled to about 45 C. are used for making plates 
and bouillon or litmus milk in flasks containing 100 c.c. are preferred for 
fluid media. The plates should be poured at once. A medium of ox-bile, 
or of ox-bile and peptone, is now considered best for B. typhosus. 

The amount of blood to be poured into each tube or flask varies some- 
what according to the type of organisms suspected to be present, from 
equal parts of blood and agar to one volume of blood in five of agar; in 
flasks, 1 to 2 c.c. of blood in 100 c.c. of medium. 

The colon group grows better in bouillon, the pneumococcus better in 
milk. Anaerobic cultures may be made in the ordinary ways. 

If after 24 hours' incubation the plates show only a few surface colonies, 
contamination certainly has occurred. Only deep colonies which appear 
similar in several or all plates should be used for subculture. True mixed 
infection in the blood is uncommon. Plates and flasks should be examined 
daily for from 5 to 14 days before discarding them as sterile. 

Value of Blood-Cultures for Diagnosis. — With the increase of laboratory 
facilities blood-cultures have become more and more important in diagnosis. 
In many instances this affords the only means of accurate diagnosis. Fol- 
lowing improvement in methods positive blood- cultures have been more 
and more frequently obtained. Formerly we were able to grow only the 
hardier organisms or those organisms in bloods which had lost some of 
their protective properties. Now certain organisms are so often obtained 



THE BLOOD 563 

that their pathological importance is doubted. Especially is this true of the 
so-called diphtheroid group. A blood-culture which contains these is now 
considered as "negative" and the same may prove to be true of some 
varieties of streptococci. In other words, the discovery of an organism in 
the blood must be evaluated since its presence may or may not be important 
in the diagnosis of the patient's present condition. 

The hardier pyogenic organisms (streptococci and staphylococci) are 
usually readily obtained in cultures in cases of general infections, osteo- 
myelitis or malignant endocarditis due to their presence. Some idea of the 
intensity of the infection may be gathered from the number of colonies 
obtained per cubic centimeter of blood used. 

The Streptococcus Group. — During the last few years different 
varieties of streptococci (see page 19) have claimed especial attention. 
For the study of these we quote Kinsella's technic. 

From 30 to 40 c.c. of blood are withdrawn from the cubital vein. From 
5 to 10 c.c. of this blood are planted directly in flasks of broth, or dextrose 
agar plates are poured, using from 2 to 3 c.c. of blood in each. Another 
5 c.c. are hemolyzed in 40 c.c. of sterile distilled water and the sediment, 
after centrifugalization, planted in a deep tube of melted ascitic -dextrose 
agar and allowed to harden before incubating. 

The cultural characteristics of the streptococci include (a) their appear- 
ance in broth, (b) their effect on serum-dextrose-agar, (c) their solubility in 
bile, (d) the lethal dose of a 24-hour broth culture for white mice, (e) their 
effect on red blood-cells (this characteristic is tested by planting cultures on 
blood-agar plates and by mixing a 24 hour broth culture with a 5% saline 
suspension of sheep red blood corpuscles (Lyall), and (/) the fermentation 
reactions on litmus milk and on rafhnose, inulin, salicin and mannite serum 
water mediums. Each strain studied should be started from a single colony. 

For the purpose of attempting a classification of the streptococci on an 
immunologic basis, rabbits are inoculated with saline emulsions of killed 
streptococci at 4-day intervals in doses equivalent to 10 c.c. of the broth 
culture. The injections are continued until the serum of an animal shows 
marked complement-binding capacity for the corresponding streptococcus 
antigen. The serum is then tested against the other strains for cross 
fixation and cross agglutination. 

Typhoid bacilli have been demonstrated in the blood in upwaid of 
75% of a series of cases by Cole, Buxton, Schotmuller, Hewlett and others, 
often days or even weeks before the Widal test is positive. 

In the paratyphoid and paracolon infections the isolation of the 
organism from the blood or stools is the only definite means of differen- 
tiating these cases from those of true typhoid fever. 

In pneumococcus infections the percentage of positive cultures is fairly 
large, the organism being found principally in the graver cases. 

Among other organisms of less frequent occurrence in the blood during 



564 CLINICAL DIAGNOSIS 

life may be mentioned: B. aerogenes capsulatus, B. coli, B. pyocyaneus, 
B. anthracis, etc. 

Blood cultures should be made if possible during a period of rising tem- 
perature and yet positive results may be obtained during the hours of 
apyrexia. 

Blood-cultures involve but little inconvenience to the patient and should 
be repeated many times before the attempt to isolate an organism is 
abandoned. 

AGGLUTINATION PHENOMENA 

Through the action on the tissues of certain bacteria soluble bodies 
appear in the blood known as agglutinins. These agglutinins, when suffi- 
ciently concentrated, have the property of clumping and rendering non- 
motile that organism whose activities gave rise to their production. 

.The nature of the interactions between the bacteria and the aggluti- 
nating serum is unknown and theoretical discussion of the phenomena 
would carry us too far afield except to say that agglutination would seem 
due to the earliest changes in that process which ends in bacteriolysis. 

Gruber-Widal Test. — This is an agglutination phenomenon applied 
to the diagnosis of typhoid fever. 

Cultures. — A standard stock culture of B. typhosus, and one which 
is actively motile, should be grown for this purpose. An organism cultivated 
through many generations on artificial media is preferred. A subculture 
on agar from this stock culture from 12 to 24 hours old is used in the test. 
Some authorities prefer fresh (10 to 18 hour old) bouillon cultures from the 
stock. Others use bouillon cultures killed by the addition of carbolic acid, 
formalin, etc. Hastings has devised a method, based on Ficker's ''Typhus 
diagnosticum," which yields very satisfactory results, viz.: To a mixture 
containing aqueous 5% carbolic acid 5 c.c, glycerin 10 c.c, sterile 0.8% 
sodium chloride solution 85 c.c. are added the growth scraped from two 24- 
hour agar slant cultures of the typhoid bacillus. Carbolic acid is preferred 
to formalin for this purpose since the latter may precipitate nocculi of pro- 
teid from the serum. The bacilli are gradually and thoroughly rubbed into 
the solution with a small spatula and this allowed to stand 5 or 6 days before 
using. The test is made by mixing equal volumes of this suspension and of 
the diluted sera. 

More satisfactory results are obtained with emulsions of the fresh living 
culture on agar (rather dry slants are best) in 0.8% salt solution or in bouil- 
lon. A loopful of the growth is rubbed against the side of the tube of salt 
solution until it is thoroughly broken up and is then gradually mixed with 
the fluid. With a little care a suspension free from clumps may be secured. 
One loopful (using a loop of standard size) in 1 c.c. of salt solution will give 
a fairly constant suspension for comparative work. 

Collecting the Blood. — Glass tubes 2 inches in length and % inch in 
diameter are drawn out into a capillary at both ends . (See Fig . 130.) 



THE BLOOD 565 

The blood is allowed to flow from a free-flowing puncture in the ear or 
finger-tip into the tube by capillary attraction until it is % full. The tube 
then lies flat until the serum has separated from the coagulum. It is then 
filed and broken off at a point just beyond the clot and the serum withdrawn 
with a capillary pipette. The separation of the serum may be hastened and 
more obtained by sealing the tips of the tube in a flame and centrif ugalizing 
it for a few minutes. This will condense the clot at one end of the tube. If 
it is desired to preserve the specimen or to send it away, both its ends maybe 
sealed in the flame or with sealing-wax. Serum is best kept in a sterile tube 
and separated from the corpuscles. If larger amounts of serum are required, 
a vein should be aspirated with the syringe (see page 561). 

Diluting the Serum. — A simple and very satisfactory method of 
diluting the serum is as follows : A piece of ^-inch glass tubing is drawn into 
a long capillary, as shown in Fig. 131. This is plunged into serum in the 
collecting tube and the capillary allowed to fill, care being taken not to 
stir up the corpuscular layer. The serum is dropped from this capillary into 
the tubes or dishes in which the dilutions are to be made. A small water- 
color porcelain palette is very convenient for making a number of dilutions, 
or salt-cellars or watch-crystals may be used. As a routine at least 2 dilu- 
tions of each serum should be made, 1 to 50 and 1 to 100. 

Using this same pipette, which has been washed out with distilled water 
to remove every trace of serum and then dried in the flame, one now adds 
to the first drop of serum 24 drops and to the second 49 drops of 0.8% salt 
solution. The addition to these dilutions of an equal volume of the sus- 
pension of the typhoid culture will give us dilutions of 1/50 and 1/100. In 
the same way any other desired dilution may be made. If greater accuracy 
or very high dilutions be desired special mixing pipettes similar to the 
Zeiss melangeur for blood counting may be employed. Again, using such 
a melangeur the whole blood may be diluted with salt solution and 
each 2 volumes of blood counted as 1 volume of serum. This mixture is 
allowed to settle or, better, is centrifugalized to remove the corpuscles and 
the diluted serum used for the macroscopic or microscopic tests. 

A. Macroscopic Method. — The macroscopic method depends on the 
agglutination and eventual precipitation of the organisms in clumps visible 
to the naked eye leaving a clear supernatant fluid. The serum is diluted 
in a small test-tube and a suspension of the organisms, living or killed, 
added; or, what is perhaps better, the full dilution of serum with salt so- 
lution, 1 in 50 or 1 in 100, is first made and the solid growth of the organisms 
then suspended in the diluted serum (see page 564). The tube is then 
examined by strong transmitted light to be sure that its contents are homo- 
geneous and free from accidental clumps. A narrow band of light from a 
lamp enclosed by a screen aids in detecting the early appearance of clumping. 
The test is considered positive if at a dilution of 1/50 or higher there is 
general clumping in 1 hour and complete precipitation leaving a clear 



566 CLINICAL DIAGNOSIS 

supernatant fluid after 24 hours. The reaction is hastened if the tubes are 
placed in the thermostat. 

This method has the advantage of simplicity in that a microscope is not 
required and that killed cultures may be used, thus obviating the. necessity 
for a thermostat and culture media. 

Several pharmaceutical laboratories in this country now sell killed cul- 
tures for the macroscopic Widal. 

B. The Microscopic Method. — The diluted serum may be mixed with 
the requisite volume of the typhoid suspension by the use of pipettes, as 
above noted, and a drop of the mixture observed on a hanging drop slide. 
Or, we may mix the 2 on the cover-slip directly. To do this we use a plati- 
num loop of stiff wire, the plane of the loop being at right angles to the 
handle and the diameter of the loop being constant. The loop is dipped 
vertically into the serum dilution and the drop so obtained placed on the 
center of the cover-slip. The loop is flamed off and dipped into the typhoid 
suspension in the same way and the 2 drops thoroughly mixed on the cover- 
slip. Approximately equal volumes are readily obtained by this simple 
method. The cover-slip is then inverted over the well of a hanging drop 
slide which previously has been ringed about with olive oil or vaseline 
and the preparation is then ready for examination. The hanging drop is 
observed with a moderately high dry lens (Zeiss D. or Leitz % in.) and by 
artificial illumination. The Argand burner or oil-lamp with a yellow flame 
is preferred. The light is stopped down with the diaphragm so as to bring 
out the refracti vity of the bacteria. 

Inspection of the freshly made hanging drop should show an absence of 
clumps and all the organisms in active motion (see Fig. 132). After the 
lapse of 1 hour if the test is positive (see Fig. 133) the organisms in a dilution 
of 1/50 will be seen to have lost their motility and to be collected entirely 
in clumps. The presence of a few non-motile free organisms in a field other- 
wise well clumped is not considered to vitiate the test. 

In such a preparation, if serum diluted 1 150 or more will in 1 hour agglu- 
tinate all the organisms into clumps, the result is positive; if the clumping 
is fair and the single organisms are non-motile this result also is considered 
positive ; but if even with fairly good clumping many of the single organisms 
are still in motion the result is considered merely suggestive. 

It is frequently noticed that the clumping is better at the higher dilutions 
and that there is marked bacteriolysis in dilutions of 1 : 10 or 1 120 or even 
higher. It also is true that many normal serums will give perfect agglu- 
tinations at 1:10 and show no trace of the reaction in a 1 150 or higher di- 
lution; therefore the tests based on the low dilutions alone are unreliable. 

The macroscopic method has rapidly gained favorin the best laboratories 
and is probably less open to error than the microscopic, provided strict 
limits of time and dilution (1 hour at dilution of 1 in 50 or higher) are ob- 
served. There is so much difference of opinion as to what constitutes micro- 



THE BLOOD 567 

scopic clumping that it is often difficult to compare results. Some authors 
consider the aggregation of a very few of the organisms to be a positive 
test. These differences of opinion have led to much confusion, particularly 
in experimental work. 

'Agglutination with Dried Blood. — If blood dried on glass, tin- 
foil, or glazed paper is to be tested the results are fairly accurate if the 
blood used is carefully weighed and the dilution based on weight 
instead of volume. 

Value of Agglutination Reactions in Typhoid Fever. — While the 
Widal reaction rarely fails to appear sooner or later in each case of typhoid 
fever it may be long delayed, even until convalescence, and it is seldom pres- 
ent before the seventh or eighth day, so that it is not an aid in early diag- 
nosis. Still, it remains our most certain confirmatory test after the bacilli 
have disappeared from the circulating blood and is indispensable in abortive, 
doubtful, and obscure cases. 

The persistence of the agglutinative reaction is variable. In some it 
remains positive for a few weeks, in others for years, after the attack of 
fever. These cases of long persistent Widal have been attributed to the 
presence of typhoid baccilli in the gall-bladder, in gall-stones, or in the 
urinary bladder. 

The agglutination of B. typhosus by normal serum diluted 1/50 in 1 
hour is so rare as to be negligible. 

The question of ''associated" or "group" agglutinations, that is, the 
agglutination of two or more closely related organisms by the same serum, as 
B. coli, B. alkaligines and B. typhosus, is too complicated to find place 
here. Suffice it so say that the limited time and the high dilution employed 
in our tests are sufficient to give us reliable specific results. 

Paracolon Infections. — While the 1 1 or more types of paratyphoid and 
paracolon bacilli often give highly specific agglutinations the presence of 
associated agglutinins should be considered and the diagnosis of any one 
type of paracolon organism by the agglutination reaction alone would be 
questionable unless confirmed by positive cultures. 

Other Agglutinations. — The agglutination reactions have been applied 
to many different organisms with more or less definite results, but in most 
cases they have not reached any considerable diagnostic value and are 
often very difficult of application. 

OPSONINS 

Phagocytosis has long been looked upon as one of the strong defenses of the body 
against infection. It has been observed that as a rule bacteria are ingested by the phago- 
cytes more readily in the presence of serum than in its absence. This action of the 
serum in facilitating phagocytosis is referred to the presence in the serum of a hypo- 
thetical " body " to which the term " opsonin " has been applied and which is supposed 
to act upon the bacteria, producing some change in them which facilitates their ingestion 
by the phagocytes. 



568 CLINICAL DIAGNOSIS 

That the action of the serum is directed toward the bacteria rather than the phago- 
cytes is indicated by the following results: Bacteria which have remained in contact 
with serum at 3 7. 5 C. for a short time and then repeatedly washed in normal salt 
solution to remove the serum are readily ingested by washed leucocytes, while bacteria 
which have not been exposed to the action of serum are ingested to a much less extent 
by washed leucocytes. This is not universally true of all varieties of bacteria. Some, 
such as Bacillus pyocyaneus, may be readily ingested by the leucocytes without previ- 
ously having been acted on by serum and a greater or less amount of phagocytosis of 
practically any variety of bacterium occurs independently of the action of serum (the 
so-called " spontaneous " phagocytosis). Spontaneous phagocytosis is said to be in- 
hibited by a 1.2% concentration of sodium chloride. Not only do different varieties of 
bacteria differ in their resistance to phagocytosis but different strains of the same 
variety may show a marked variation in their resistance to phagocytosis even under 
the influence of the same serum. In general the more virulent the strain the more 
resistant it is to phagocytosis. 

Opsonins are placed in the category of immune bodies along with agglutinins, 
precipitins, bacteriolysins, etc., and like them are regarded as specific bodies; that is, 
just as the agglutinins for different varieties of bacteria are specific so there are specific 
opsonins. Like the agglutinins also, opsonins occur normally in the serum and in greater 
amounts as a result of infection or immunization. The former are designated as "normal" 
opsonins, the latter as " immune " opsonins. Normal opsonins are said to be thermo- 
labile, being destroyed by an exposure to 57 C. for %, hour, while immune opsonins 
resist this exposure and are therefore thermo-stabile. 

If the foregoing observations are correct and if opsonins are really specific immune 
bodies playing a very important part in the defensive mechanism of the body, any 
method which would enable their accurate estimation might be of great service in diag- 
nosis and prognosis and any means of regulating their presence in the body might be of 
great therapeutic importance. 

For diagnostic purposes the estimation of any of the other immune bodies — agglu- 
tinins, precipitins, complement-fixing amboceptors, etc. — is at present more valu- 
able than the opsonic index. Prognostically little value can be attached to any of 
them and by therapeutic measures these other bodies can be influenced more than can 
the opsonins. 

The power of the serum in favoring phagocytosis is spoken of as its opsonic power. 
The opsonic index of a given individual is the ratio of the opsonic power of his serum 
to the opsonic power of the serum of a normal individual. 

Since the opsonic index is so little used now, although the reason may be our faulty 
methods of technic, we will not give a detailed description of the technic, merely enough 
that the student may have a general idea of a test so often referred to in literature. 

Equal quantities of the patient's serum, of a suspension of washed leucocytes ob- 
tained from any source, and of a suspension of the bacteria to be tested are mixed and 
incubated at 3 7. 5 C. Smears are then made from the mixture and after appropriate 
staining the average number of bacteria ingested per leucocyte js determined. This 
number represents the opsonic power of the serum and is sometimes spoken of as the 
phagocytic index. At the same time a similar specimen is prepared, only the serum 
used is that of a person known to be normal so far as that organism is concerned. 

The ratio which the phagocytic index of a given serum bears to the phagocytic index 
of a normal serum is the opsonic index. 

For example: Suppose the average number of bacteria taken up by the leucocytes 
in a preparation in which patient's serum has been used is 3, and the average number 
taken up by the leucocytes in a preparation in which normal serum has been used is 6, 
the ratio would be 3 : 6, and the opsonic index 0.5. Had the average number in the 
preparation in which patient's serum was used been 9 the opsonic index would be 1.5. 



THE BLOOD 569 

COMPLEMENT FIXATION 

The principles which underlie the complement fixation tests are those 
first demonstrated by Bordet and Gengou, viz., that if antigen and its 
specific amboceptor are brought together in the presence of complement, the 
complement enters into combination with them ; and if the antigen and am- 
boceptor are present in sufficient amounts the complement is bound, or 
used up, in the combination. Evidence of this reaction is seen in all the 
cytolytic, bacteriolytic and proteolytic immune reactions. The part played 
by complement is probably that of an enzyme which acts upon the antigen 
causing its lysis provided a sensitizer specific to that antigen is present. 
Ehrlich's nomenclature of his third order of antibodies is used in the fol- 
lowing discussions but this does not imply an acceptance of his conception 
of the mechanism of the reaction. This reaction of these 3 elements is so 
constant that any 2 of them may be used as a qualitative bio-chemical test 
to determine the presence or absence of the third. For example, given an- 
tigen (which may be any protein, cellular or in solution) and complement, 
and the presence of amboceptor specific to that given antigen may be de- 
tected. Given a specific amboceptor and complement and the presence of 
the antigen for which that particular amboceptor is specific may be detected. 
Given an antigen and its specific amboceptor and the presence of comple- 
ment may be detected. In this way sensitive specific tests may be applied 
to the detection of meat adulteration, the character of blood stains, the 
identification of bacteria, etc., as well as to serological diagnosis. Thus, 
as originally shown by Bordet and Gengou, if an extract of typhoid bacilli 
and blood serum from a typhoid patient are brought together in proper 
proportions and incubated for a short time at body temperature, the com- 
plement in the serum, sensitized by the presence of typhoid amboceptor in 
the serum, will unite with the typhoid extract. If typhoid amboceptor is 
not present in this patient's serum, such union will not occur and the 
complement will remain free. Since it would not be possible to detect 
optically whether the complement remains free or becomes bound an 
indicator is necessary. 

An indicator often used consists of washed sheep's erythrocytes and 
the serum of a rabbit which has been immunized against sheep corpuscles. 
If a suitable amount of this indicator is added after incubation to the tube 
containing the patient's serum and typhoid extract the presence of free com- 
plement would be indicated by hemolysis, i.e., laking of the sheep corpuscles. 
Such a reaction would show that this particular patient's serum contains no 
specific amboceptor for typhoid bacilli to bind the complement to the anti- 
gen, i. e.y to the typhoid extract. If the patient's serum does contain am- 
boceptor for typhoid bacilli, they together would bind or absorb the com- 
plement in their combination and then when the indicator is added no 
hemolysis, i. e., laking, of the corpuscles would occur. The hemolysis is 
easily noted optically, and when the quantities of the reagents are properly 



570 CLINICAL DIAGNOSIS 

adjusted this test may be made very delicate. Too great emphasis cannot 
be laid upon the quantitative adjustment of the reagents since the reliability 
and delicacy of all complement fixation depend very directly on this. 

Glassware and Material. — The test tubes recommended .for sero- 
logical work measure 7 cm. long and 1 cm. in diameter. These hold ap- 
proximately 6 c.c. Other sizes may be used if desired. Mohr pipettes of 
the following sizes are desirable: A 5 c.c. pipette graduated in 0.1'c.c, a 
2 c.c. pipette graduated in 0.1 c.c; a 1 c.c. pipette graduated in 0.01 c.c; 
and one containing 0.2 c.c. graduated in 0.0 1 c.c. It is desirable that the 
graduations should in each case extend to the tip of the pipette. Graduated 
cylinders of 25, 50 and 100 c.c capacity for making up solutions and glass 
beakers of 50 or roo c.c. capacity are necessary. • 

The most convenient test tube rack is made of metal and holds a double 
row of 10 tubes each. The water bath should have a thermo-regulator 
capable of maintaining a constant temperature of 38°C. All glassware used 
should be reserved for this work only and should be kept scrupulously clean. 
After use the tubes and pipettes should immediately be emptied, rinsed in 
tap water, then either boiled or allowed to stand several hours in distilled 
water. Then the tubes may be inverted in a wire basket until dry. It is 
our practice to sterilize the glassware with dry heat as a final step in prep- 
aration, though bacteriological asepsis is not necessary. Any glassware 
which becomes permanently clouded should be discarded. Especial care 
must be used to wash every trace of reagents as well as of chemical cleaning 
solution from the pipettes before drying them. 

Preparation of Hemolytic Amboceptor. — Because of their use as 
indicators in all complement fixation tests hemolysins have a great practical 
value. They are produced by injecting into one animal the red corpuscles of 
an animal of another species. Rabbits are the animals most commonly used 
for this purpose. They vary widely in their power to form hemolysins, but 
form a very potent hemolytic amboceptor when injected with sheep's 
corpuscles. Human blood, beef blood or that of fowls may be used. In 
many laboratories human blood is used because it is so easily obtained, 
yet it is more difficult to produce powerful hemolysins with human than with 
sheep corpuscles. 

The sheep's blood may be obtained from the jugular vein of the animal 
by means of syringe, or it may be secured from an abattoir. It should be 
received into a clean sterile flask containing glass beads and shaken vig- 
orously for 5 minutes to remove the fibrin, or it may be received into an 
equal quantity of isotonic salt solution containing 0.5% of sodium citrate to 
prevent clotting, then centrifugalized and the serum drawn off. The cor- 
puscles are next freed of all serum by washing and centrifugalizing them 3 
times, using each time from 5 to 10 times their volume of physiological 
salt solution. 

Hemolysins appear in the rabbit's blood following the intra-peritoneal 



THE BLOOD 571 

or intravenous injections of the washed corpuscles. Aseptic technic 
should be employed throughout. It is advisable to filter the corpuscle 
suspension in order to remove small clots of fibrin. The potency of the 
amboceptor produced bears no direct relation to the size of the doses of 
corpuscles injected ; for example, a highly potent hemolytic serum may 
be prepared by three intravenous injections at intervals of 3 days of 

3 c.c., 5 c.c, or 7 c.c. of a 10% suspension of sheep corpuscles. After 
from 6 to 10 days following the last injection a specimen of blood should be 
obtained from the rabbit's ear and the serum tested for its hemolytic power 
(see below). 

Some, using a slower method, inject larger doses of the sheep's corpuscle 
and at longer intervals. For example, they inject 3 c.c, 5 c.c, 10 c.c, 15 c.c, 
and 20 c.c respectively of a 10% corpuscle suspension at intervals of 6 or 
7 days. 

Instead of injecting the corpuscles intravenously the same quantities 
may be introduced intra -peritoneally at the same intervals. This method 
has the advantage that the animals are not so apt to die from anaphylaxis 
as when the intravenous method is used. 

In the preparation of anti-human amboceptor Noguchi advises to give 

4 injections of 4 c.c, 3 c.c, 4 c.c, 3 c.c, and possibly another of 4 c.c, at 
intervals of 4 or 5 days. 

In rmmunizing rabbits against corpuscles it is advisable to use 2 or more 
animals simultaneously, as it frequently happens that only 1 of several will 
produce a highly potent serum. 

One week after the last injection about 1 c.c. of blood is drawn from the 
ear of the rabbit and its serum titrated to determine its hemolytic strength. 
This may be done as follows : 

In 1 row of clean test tubes is made a series of dilutions of the rabbit's 
serum with isotonic salt solution; e. g., 1:25, 1:50, 1:75, 1:100, 1:200, 1:300, 
1:400, 1:500, etc. Next, 0.1 c.c. from each of these dilutions is carefully 
measured into a corresponding row of empty tubes. This row then con- 
tains 1/250 c.c, 1/500 c.c, 1/750 c.c, 1/1000 c.c, etc., of the original rabbit 
serum. Two units of standardized (seepage 573) complement are now added 
to each tube. If none is available 0.02 c.c of guinea pig serum may be as- 
sumed to contain about 2 units of complement and this quantity used in 
testing the hemolytic serum. These small quantities are most easily ob- 
tained by diluting the serum 1:15 and then measuring o. 1 c.c. into each tube, 
or by diluting it 1:10 and adding 0.2 c.c. into each tube. Next 0.5 c.c of a 
2 % suspension of washed sheep corpuscles (page 573) are added to each tube, 
and sufficient salt solution to make the total volume in each exactly 1.0 c.c. 
This is done that the reagents may act in uniform volumes. The set of 
tubes is now incubated for 30 minutes in a water bath at from 37°C. to 
4o°C. temperature, and the amount of hemolysis in each tube then 
noted. The smallest quantity of rabbit serum in this series which will 



572 CLINICAL DIAGNOSIS 

cause complete hemolysis is accepted as i unit of amboceptor and twice that 
amount is the dose of this serum to be used in all hemolytic work. For ex- 
ample, if hemolysis is complete in all of the first -6 tubes in the above series 
and only partial in the others, then 1/3000 c.c. is the unit of amboceptor, 
since the 6th tube contained 0.1 c.c. of rabbit serum diluted 1:300, i. e., 
1/3000 c.c. of the original serum. It very frequently happens that the 
hemolytic amboceptor is so very potent that the above series of 
dilutions will have to be extended in order to determine the potency of the 
serum. Amboceptor whose unit is 1/50,000 c.c. or less is occasionally pro- 
duced. It is our practice to discard amboceptor whose unit is a larger 
amount than .001 c.c. If the test shows the serum to be of satisfactory 
hemolytic potency, the rabbit is bled to death under aseptic precautions and 
the serum separated. It is our practice to introduce a No. 20 needle attached 
to a 30 c.c. syringe into the heart of the anesthetized animal since it is 
easier to secure the blood free from bacterial contamination by this method 
than by opening the carotid artery. 

The rabbit serum may be preserved in small sterile tubes or ampullae 
hermetically sealed, or dried on the filter paper. The most satisfactory 
method is to mix it with an equal quantity of glycerine to protect it against 
bacterial growth. This does not in any way affect the serological proper- 
ties, for the serum will retain its potency for many months if kept in a cool 
place protected from light. 

The Preparation and Standardization of Complement. — Com- 
plement would seem to be a proteolytic enzyme which is normally present 
in the serum of the mammalia. Its strength or quantity differs in different 
individuals and in different species. It is destroyed easily by heat, by 
chemical substances such as acids, alcohol, etc., and by bacterial growth. 
It rapidly becomes weaker and finally disappears if the serum is allowed to 
stand exposed to warmth and light. 

According to some methods of complement fixation use is made of the 
complement present in the patient's own serum. This is open to the objec- 
tions that its amount varies so widely in different individuals and in the 
same person in disease conditions that to determine the strength of each 
patient's own complement at the time we planned to use it would involve 
much unnecessary work. 

Complement is most active and least variable in amount in the serum of 
guinea pigs, hence this is the accepted source. Only vigorous healthy 
animals should be used, and especially animals which have not been used 
for any other purposes. It is advisable to use the pooled serum of 2 or more 
animals. The blood may be obtained either by opening the carotid artery 
and bleeding the animal to death or by introducing a No. 22 needle directly 
into its heart and withdrawing 4 or 5 c.c. of blood into a syringe. The latter 
method has the advantage of economy of animals since they may be bled 
repeatedly at intervals of 2 or 3 weeks. The blood is allowed to stand in a 



THE BLOOD 573 

perfectly clean tube at room temperature until firmly clotted, the clot then 
loosened from the walls of the tube and the tube then placed in an ice box 
while the serum is allowed to separate. Or, the serum may be separated by 
centrifugation immediately after drawing the blood. The serum should 
be carefully removed from the clot or sediment so that it may be quite 
free from cells and hemoglobin. 

In all work which involves the complementary property of animal serum 
it is very important that definite amounts of complement be used. Careful 
titration of the complement is therefore a most important preliminary step. 
The unit of complement is the smallest quantity which will produce complete 
hemolysis of the standard unit of blood cells mixed with two units of hemo- 
lytic amboceptor. It therefore depends directly upon the quantity of cor- 
puscles used in the hemolytic system. 

In the test as originally performed by Wassermann i.o c.c. of a 5% 
suspension of sheep corpuscles was the unit, or TV, used. This was soon 
reduced to y% that quantity (o. 5 iV) , while now it is our practice to use o. 5 c.c. 
of a 2% suspension (0.2 N) of washed sheep corpuscles; that is, the equiva- 
lent of 1.0 c.c. of a 1% suspension. That is, the unit of complement we 
use is % the amount originally used and % of that recommended by many 
serologists. The advantage of a small unit of complement is that it greatly 
increases the delicacy of the test. Often the specific antibody for which 
we are testing is present in the patient's serum in very small amounts. If, 
therefore, there is scarcely enough antibody present to bind a quantity of 
complement represented by 0.2 N in the presence of the appropriate antigen, 
there would obviously be insufficient antibody to bind the amount repre- 
sented by 0.5 N or N. Therefore a definitely positive reaction with the 
smaller unit of complement would be found negative if tested by a system in 
which a larger unit of complement was used. 

The hemolytic system consists of complement, red blood-cells and ambo- 
ceptor specific to those cells. Both the amboceptor and the complement are 
substances which vary in strength. The amount of blood corpuscles is the 
only easily controlled factor and hence is the unit to which the hemolytic 
system is adjusted. In order to provide a corpuscle suspension of as nearly 
constant value as possible, we add about 5 c.c. of sheep blood to about 45 
c.c. of 0.85% salt solution, mix thoroughly and centrifugalize. This washing 
and centrifugation are repeated twice. In the centrifugation both speed and 
time are controlled, i. e., 2000 revolutions per minute for 15 minutes. This 
will give a corpuscle sediment of almost constant density. One cubic 
centimeter of this sediment is carefully measured and made up to 50 c.c. 
with salt solution. This gives a fairly accurate 2% corpuscle suspension 
with which both complement and amboceptor may be standardized . 

Titration of Complement. — A small amount of fresh guinea pig serum, 
obtained as described above, is diluted 1:20 with salt solution. Into a 
series of clean test tubes the following quantities of diluted complement are 



574 CLINICAL DIAGNOSIS 

carefully measured, using a i c.c. pipette graduated in hundredths: o.i c.c., 
0.15 c.c., 0.2 c.c., 0.25 c.c., 0.3 c.c., 0.35 c.c., 0.4 c.c., etc. To each tube are 
then added 2 units of hemolytic amboceptor, determined as described on 
page 570, and 0.5 c.c. of 2% washed corpuscle suspension. A sufficient 
quantity of salt solution is now added to bring the total volume in the 
tube up to 1.0 c.c. The set is then incubated in a water bath for 30 min- 
utes after which time the amount of hemolysis in each tube is noted. The 
smallest quantity of complement which produced complete hemolysis of 
the corpuscles is accepted as one unit of complement. Thus, in the above 
series, if hemolysis was only partial in the first 2 tubes and complete in the 
third and in all the remaining tubes, the unit of complement would be the 
amount present in the third tube, which is 0.2 c.c. of complement diluted 
1:20, or o. 01 c.c. of undiluted complement. 

Those who prefer the larger unit of corpuscles (see above) make the 
dilutions of complement as above, but add 0.5 c.c. of a 5% suspension of 
corpuscles and 2 units of amboceptor. 

Whatever hemolytic system is used the density of the corpuscle suspen- 
sion should be maintained as uniform as possible and that exact minimal 
amount of complement and amboceptor determined which will produce 
complete hemolysis of the unit of corpuscles used. 

The Patient's Serum.— The serum is obtained by introducing a needle 
into a superficial vein and drawing the blood directly into a glass syringe. 
The vein usually chosen is the median cephalic on the anterior surface of 
the arm. The skin is sterilized by soap and water followed by an alcohol and 
iodine solution, or by Harrington's solution. The needle should have a 
caliber not larger than No. 2 1 and should be sterile. It is essential that the 
syringe be absolutely clean, dry and free from any trace of chemical sub- 
stance. For this reason the syringe should not be washed with alcohol. A 
rubber tourniquet is applied above the elbow sufficiently tight to produce 
distention of the veins. The needle is inserted with the shank almost 
parallel to the skin surface, and with flat side of the point up. From 2 c.c. 
to 5 c.c. of blood is drawn into the syringe, the tourniquet then removed and 
the needle withdrawn. Pressure is made at the point of puncture with a 
sterile sponge or cotton immediately as the needle is withdrawn. The blood 
is transferred at once to a clean, dry tube and allowed to stand at room 
temperature until clotted. The clot is then separated from the sides of the 
tube and the specimen placed in a refrigerator for several hours. Usually a 
sufficient quantity of clear serum can be pipetted directly from the tube; 
if not, the specimen may be centrifugalized and the clear serum drawn off. 

Before testing the serum it is necessary to destroy all of its comple- 
mentary property. While many recommend that the serum be heated for 
30 minutes at 5 6° C. we have found that heating it for even 5 minutes at 
55 C. will completely inactivate the complement and hence have adopted 
the routine of heating the serum for 10 minutes at 55° to 56 C. In this 



THE BLOOD 575 

way we avoid changing by the prolonged heating other properties of the 
serum than the complement. 

It should be the rule to draw the patient's blood for serological tests be- 
fore meals, or at least several hours after the last meal, since the products of 
recent digestion may render the serum anti-complementary. The presence 
of water, alcohol or other chemical in the glassware used may have a similar 
effect, or may produce sufficient hemolysis to make the serum unstable. 
The growth of bacteria in serum frequently produces anti-complementary 
properties and sometimes false positive reactions. For these reasons strict 
cleanliness should be observed in handling the serum and the tests should be 
made as soon as possible after the blood is drawn. It should be an invariable 
rule to keep all serological reagents in a refrigerator at a temperature slightly 
above o° C. until they are used. 

The Wassermann Test. — The antigen-amboceptor-complement 108 
reaction was at first used for the detection of antibodies in sera and for the 
identification of bacteria. It was found to be exquisitely delicate, detecting 
minute amounts of antigens with the sharpest specificity limits of all the 
immunity reactions. It can even be used to determine the presence in 
tissues of specific organisms which cannot be cultivated; e. g., the presence 
of a specific scarlatinal virus in the tissues of a patient with this disease. 
This fact led Wassermann, using as antigen extracts of the livers of con- 
genital syphilitic fetuses, which contain great quantities of spirochetes, to 
attempt with this test to determine in a given serum the presence of specific 
amboceptors for the virus of syphilis, such amboceptors being present in 
persons infected with syphilis as a result of their reaction to the infection. 
As originally introduced, then, the Wassermann reaction was supposed to 
be a specific reaction, similar to the original complement fixation reaction of 
Bordet and Gengou between syphilitic antigen, specific syphilitic ambo- 
ceptors and non-specific complement. It was soon discovered, however, 
that the reaction as it occurred in syphilis was decidedly different, since in 
place of a specific antigen (the extracts of tissues containing the syphilitic 
virus) it is possible to substitute the most varied sorts of tissue extracts 
which certainly are free from spirochetes (e_. g., ox heart). Now, syphilitic 
tissues are seldom used. One may even use with fairly satisfactory results 
commercial lecithin or mixtures of commercial lecithin and sodium oleate. 
Noguchi and Bronfenbrenner conclude as follows : ' ' We know merely this : 
that complement in the presence of syphilitic antigen may be rendered 
inactive by one or more substances in the body fluids of a syphilitic or 
parasyphilitic patient. " 

Extended investigation of these non-specific "antigens" which give 
specific complement fixation with syphilitic sera has shown them to be 
related to the lipoids, especially the lecithins, as indicated by the fact that 

106 Modified from Wells, Chem. Path., pp. 235-237. 



576 CLINICAL DIAGNOSIS 

the most efficient "antigens" contain the acetone-insoluble fraction ot the 
tissue lipoids. The antigenic value of this fraction of different liver extracts 
varies almost directly with its power to combine with iodine (Noguchi and 
Bronfenbrenner) , which would indicate that unsaturated fatty acids are 
important in the reaction. Lecithins from different sources vary in effi- 
ciency ; heart lecithin is more active than liver lecithin while brain and egg 
yolk lecithin follow. The addition of cholesterol to the lecithin solutions 
greatly increases their activity. An acetone-precipitated "antigen" of this 
class certainly is not a true antigen, for fixation antibodies are not developed 
in animals injected with those very lipoids which have been shown to be 
quite efficient in the Wassermann reaction. 

The substance in the syphilitic serum which participates in the 
Wassermann reaction would seem to be related to the globulins, 
especially euglobulin, which are decidedly increased in the blood and 
spinal fluid of syphilitics. 

P. Schmidt explains the reaction as due to the physico-chemical properties of the 
globulins of the syphilitic serum, which, he believes, possess a greater affinity for the 
colloids of the antigen than do normal globulins; this affinity is held in check in normal 
serum by the albumins of the serum, which in lues are relatively or absolutely decreased. 

A favorite interpretation of the Wassermann reaction, which seems to harmonize 
with the known facts, is that there is a precipitation of serum globulin by the lipoidal 
colloids of the antigen and absorption of the complement by this precipitate. 

Wassermann 108 and his collaborators also used salt-solution extracts 
chiefly of the spleen of a syphilitic fetus. The tissue, was cut into small 
pieces. To i part by weight of this substance were added 4 parts of normal 
salt solution and 0.5% of carbolic acid. This was shaken in a shaking ap- 
paratus for 24 hours and then the coarser particles removed by centrifu- 
gation. The reddish supernatant fluid was used as the antigen. This could 
be preserved for a long time in dark bottles in the ice chest. Alcoholic 
extracts of syphilitic organs were later used by several authors who ex- 
tracted syphilitic liver tissue for 24 hours with 5 times its volume of absolute 
alcohol. This was filtered through paper and the alcohol evaporated in 
vacuo at a temperature not above 40 C. About 1 gram of this material was 
then emulsified in 100 c.c. of salt solution to which 0.5% of carbolic acid had 
been added. 

The Antigen. — While the term "antigen" as ordinarily used in the 
Wassermann reaction is, strictly speaking, a misnomer (see above) yet it is 
so generally used that it may be retained, but with a distinct understanding 
as to its actual meaning. 

It is also of interest that in practical work this artificial antigen is even 
superior to the real antigen since the pure Treponema pallidum extract gives 
reactions in only a few late tertiary cases and results which run not at all 
parallel to the fixations obtained with non-specific lipoidal substances. Al- 

108 Modified from Hiss and Zinsser, pp. 263-264. 



THE BLOOD 577 

though we are, at the present writing, still in the dark as to whether the 
syphilitic antigen depends for its properties upon the lipoidal nature of the 
extracts, or upon the size and dispersion of the particles present in the ex- 
tracts, we can still assert that the test is reliable and, with care in execution 
and interpretation, is of enormous value in the diagnosis of syphilis. It is 
necessary, however, to recognize that it surely is not a specific antigen- 
antibody reaction. 

The antigens in common use to-day are prepared as follows : 

1. Beef, sheep, human or guinea-pig heart muscle finely chopped up and extracted 
in 5 times its volume of absolute alcohol. This mixture is kept for from 5 to 7 days 
in the incubator, during which' time it is frequently shaken. It is then filtered 
and titrated. This preparation of antigen is known as " plain alcoholic extract of 
heart muscle." 

2. Noguchi's Acetone-Insoluble Lipoid Antigen. — Fresh spleen or heart muscle is 
macerated and extracted for from 5 to 7 days in the incubator in 5 times its volume of 
absolute alcohol, during which time it is frequently shaken. It is then filtered and 
evaporated to dryness with the aid of a fan. The sticky residue is taken up in a small 
quantity of ether and this ether solution poured into 4 times its volume of acetone (C. P.). 
The floccular precipitate which forms is collected and can be preserved under acetone. 
About 0.2 gms. of this paste is dissolved in 1 c.c. of ether and, added to 9 c.c. of pure 
methyl alcohol for permanent preservation. It is diluted with salt solution before using 

3. Cholesterinized Antigen. — According to the researches of Sachs and Rondoni, 
Browning and Cruikshank, and Walker and Swift antigen can be made more 
delicate by the addition of cholesterin. Walker and Swift recommend that to an alco- 
holic extract of human or guinea-pig heart enough cholesterin be added to make a 
concentration of 0.4%. 

Whichever antigen is used it must be tested and known to conform to 
the following requirements: 

It must show no anticomplementary nor hemolytic action when used 
in small amounts. 

It must be sensitive to the presence of syphilitic antibodies, as shown 
by the absorption of complement when mixed with the serum from known 
syphilitic patients. 

It must not absorb complement when mixed with non-syphilitic serum. 

There must be a wide margin between the antigenic dose and the amount 
ot the antigen which by itself would manifest anticomplementary or 
hemolytic action. 

It must keep well and develop no variations in the above characteristics. 

Different extracts prepared after the same manner should be quite 
similar in their characteristics. 

The suitability of the antigen is determined as follows: 

To estimate the degree of its anticomplementary action the following quantities 
of the antigen are carefully measured into a series of clean, dry tubes: 0.25 c.c, 0.1 c.c, 
0.075 c.c, 0.05 c.c, 0.025 c.c, 0.01 c.c, 0.0075 c.c, 0.005 c.c Two units of complement 
determined as described on page 573 are added to each of these tubes and also to a 
control tube containing no antigen. The total quantity in each tube is made up to 0.5 c.c. 

37 



578 CLINICAL DIAGNOSIS 

with normal salt solution. These tubes are incubated for 30 minutes at 38 C, then 
2 units of amboceptor and 1 unit of washed corpuscles appropriate to the amboceptor 
are added to each tube and the tubes are again incubated for 30 minutes. If in the control 
tube there is complete hemolysis and there is failure of hemolysis in the remaining tubes, 
then this antigen certainly has the property of inhibiting the action of complement. 
If there is no hemolysis in the first 2 tubes, partial in the third and complete hemolysis 
in the remaining tubes, then this antigen is anticomplementary in quantities of 0.075 
c.c. or more. 

To determine whether the antigen itself is capable of causing hemolysis, a series 
of tubes is arranged which contain the same quantities of antigen as above. No comple- 
ment is added to any of the tubes in this case. The volume of each tube is now made 
up to 0.5 c.c. with salt solution and a unit of corpuscle suspension added to each tube. 
The tubes are now incubated for 30 minutes and the results read as before. Any hemol- 
ysis noted in this series indicates that the antigen possesses hemolytic property and if 
it is present in the tubes containing the small quantities of antigen, this antigen must 
be discarded as unsatisfactory. 

To determine the sensitivity or antigenic property of the antigen the following 
quantities of antigen are carefully measured into a series of clean dry tubes: 0.05 c.c, 
0.025 c - c -» °- 01 c - c -> °- 00 75 c - c -> 0.005 c - c -> 0.0025 c.c. Sufficient salt solution is added to 
each to make the total amount 0.3 c.c. To each of these tubes is next added 0.1 c.c. 
of inactivated patient's serum which has previously been tested and is known to give 
a positive test, also 2 units of complement determined as previously described. The 
following controls also are prepared: One tube containing the patient's serum, 2 units 
of complement and sufficient salt solution to make the total volume 0.5 c.c. and 1 tube 
containing 2 units of complement alone and made up to 0.5 c.c. with salt solution. 
These tubes are incubated in a water bath for an hour at 38 C. Then 2 units of ambo- 
ceptor and 1 unit of corpuscle suspension are added to each tube and they are incubated 
again for 30 minutes. Should there be no hemolysis in the first 4 tubes and partial or 
complete in the others it would indicate that the antigen is sensitive, since 0.01 c.c. 
will bind complement in the presence of syphilitic serum. If, as in this illustration, the 
amount of the antigen which will bind complement is less than % the quantity which is 
either anticomplementary or hemolytic then the antigen is satisfactory. It usually 
is the case that satisfactory antigen is sensitive in smaller quantities than illustrated 
in this instance. Antigen should never be used whose antigenic dose approaches the 
anticomplementary or the hemolytic quantity by a narrower margin than that illus- 
trated in the above example. 

It is well to run each new antigen which has been prepared and tested 
as above described in parallel tests with an antigen previously used and 
known to be sensitive on a rather large series of routine cases before 
adopting the new antigen for routine work. 

The quantity of antigen used in routine work should not exceed twice 
the minimum sensitive quantity determined as above since it has been 
found that occasionally antigen in excess is not as sensitive as is the min- 
imum quantity. 

It is the custom in many laboratories always to use 2 or more antigens in 
routine work. If only 1 antigen is used, we prefer the acetone-insoluble frac- 
tion of the alcoholic extract of beef heart as prepared by Noguchi , since we have 
found that this usually is more delicate than plain alcoholic organ extracts. 

The cholesterinized antigen, recommended by many workers because 
it is so sensitive, may be too sensitive since the blood of individuals who 



THE BLOOD 579 

seem clinically quite free of syphilis has given positive tests with this antigen 
and negative to all other used. It is the opinion of many workers that the 
cholesterinized antigen is most useful in tests which are intended to control 
the results of the treatment of cases known to have syphilis. If it is used as 
an antigen in routine diagnostic work, other tests using another antigen 
should be run in parallel with it. For illustration, if a case gives a weakly 
positive reaction with alcoholic extract or acetone-insoluble antigen, and 
a strongly positive one with the cholesterinized antigen then the suspicion 
of syphilis is strengthened. 

Technic of the Wassermann Test. — Since the technic originally used by 
Wassermann is the basis of all modifications now in general use his method of 
performing it will be briefly outlined. Wassermann used as antigen an 
aqueous extract of the liver of cases of congenital syphilis. This antigen is 
rarely used now and any one of the antigens described above may be sub- 
stituted for it. His reagents were prepared as above described except that 
the dose of hemolytic amboceptor was determined for a larger quantity of 
sheep cells. As many pairs of tubes were placed in a rack with a double row 
of holes as there were patients' sera to be tested, each pair consisting of i 
tube in the front row and the one just behind it. Three additional pairs were 
added for controls. Into all the tubes was measured 0.7 c.c. of salt solution 
and then into each pair 0.2 c.c. of an inactivated patient's serum. This was 
done with, each of the sera to be tested and also with a serum known to be 
positive and one known to be negative. Into each tube, front and back, was 
placed 0.1 c.c. of fresh guinea pig serum; then 1 c.c. of antigen so diluted 
that this quantity contained twice its minimal antigenic dose (see page 578). 
The additional tube in the front row contained antigen and complement 
alone and served as an antigen control. The corresponding tube in the rear 
row contained only complement and served as a control on the hemolytic 
system. i The volume in each tube was now brought to a total of 3 c.c. by the 
addition of salt solution and the tubes all incubated at body temperature 
for 1 hour. Following the incubation 1 c.c. of amboceptor, so diluted that 
this quantity contained 2 hemolytic units, and 1 c.c. of a 5% suspension of 
washed sheep corpuscles were added to each tube. After another incuba- 
tion period of 1 hour the results were read. It was the practice in some 
laboratories to place the racks in an ice box for 2 or 3 hours after the second 
incubation period. This will facilitate the estimation of the hemolysis, 
particularly in tubes where it may be only partial, since on standing the 
non-hemolysed corpuscles will settle to the bottom of the tubes. 

A widely used modification of the above technic consists in using the reagents in 
exactly % the quantities specified above. 

The method we now are to describe may be regarded as the original 
Wassermann technic except that }{ the quantities of that technic are 
used. The essential variations are the extreme care used in adjusting the 



580 CLINICAL DIAGNOSIS 

hemolytic system, the fact that yi the original quantity of the patient's 
serum is used and }{ the quantity of the other reagents. This increases 
the proportion of patient's serum to the other reagents and so tends to 
increase the delicacy of the test. We believe that this is the most satis- 
factory for routine use of all of the modifications cf the Wassermann test. 
When carefully performed it combines a high degree of delicacy with a 
relatively simple technic. It is adapted to all complement fixation tests, 
as well as to that for syphilis. The directions given above for titrating the 
various reagents are intended for this modification alone. If other modifi- 
cations are used the same scheme of titration may be employed, only one 
makes suitable changes in total volumes to adapt the quantities to those of 
the modification in question. 

The antigen, complement, amboceptor and washed sheep corpuscles are 
prepared and titrated as described on the preceding pages. The patient's 
serum, free from red blood corpuscles, is inactivated at 55 for 10 minutes. 
As many pairs of tubes are placed in the rack as there are sera to be tested. 
If several, time is saved in pipetting and greater accuracy in measurement 
is obtained if the antigen, complement and salt solution are mixed together 
in suitable proportions and the proper quantity of this measured into each 
tube. For illustration, if 16 sera are to be tested, an antigen-complement 
mixture for 20 (allowing for controls) is made as follows: Assuming for 
example that the unit of complement (see page 574) is .0075 c.c, since 2 
units are used in each test then 40 such units, that is 0.3 c.c, is measured 
carefully into a dry, clean, small graduate . Assuming further that the unit of 
antigen (see page 578) is o. 01 c.c, since 2 units are used in each test then 40 
units, or 0.4 c.c, is measured accurately into 5 or 6 c.c of salt solution and 
added to the 0.3 c.c of complement. The total volume of complement anti- 
gen mixture is now brought to exactly 8 c.c with salt solution and the whole 
well mixed. Then 0.4 c.c. of this complement-antigen mixture is measured 
carefully into each of the twenty tubes of the front row in the racks. 
Diluted complement without antigen is prepared in the same proportion as 
above and 0.4 c.c. pipetted into each tube in the rear row. A 2 c.c. or a 
5 c.c pipette graduated in tenths is convenient for this measurement. 

The "set-up" is now ready to receive the sera to be tested. One tenth 
of a cubic centimeter of each serum is added to 1 tube in the front row and 
to the corresponding tube in the rear row. Then the pipette is rinsed with 
salt solution and the same amount of another serum added to each of the 
next pair. To the seventeenth pair of tubes is added the same amount of a 
serum which a previous test has shown to be definitely positive and to the 
eighteenth, a serum known to be negative. This leaves 2 pairs of tubes to 
which no human serum is added for controls of the reagents, the front tubes 
for complement-antigen and the rear tubes for complement. To each of 
these controls one adds 0.1 c.c of salt solution (in place of a serum) in 
order to make the volume in all tubes equal. 



THE BLOOD 581 

The "set-up" is now incubated in a water bath for an hour at 3 8° C, 
after which time sheep cells and their specific amboceptor are added as a 
test for the presence of free complement. Two units of amboceptor (see 
page 571) and 0.5 c.c. of 2% corpuscle suspension are added to each of all 
the tubes. Time may be saved if enough corpuscles and amboceptor for all 
are first mixed. That is, 0.5 c.c. of corpuscles from the centrifuge tube (see 
page 573) are measured with a wet pipette into 24.5 c.c. of salt solution. 
This amount is sufficient for 50 tubes. Since each tube must receive 2 
units of amboceptor 100 units of this are added to the corpuscle suspension. 
For example, if the titration of amboceptor as described on page 571 shows 
that .0015 c.c. is the unit, then 100 times this amount, or 0.15 c.c. of un- 
diluted amboceptor is added to the sheep corpuscles and thoroughly mixed. 
Then 0.5 c.c. of this mixture is measured into each tube and they all are 
again incubated. Most authors specify 1 hour for this last incubation time, 
but if the reagents are satisfactory hemolysis is complete before 30 minutes 
and further incubation produces little change. 

In performing Wassermann tests on cerebrospinal fluid it is not nec- 
essary to inactivate this fluid since it contains no complement. A larger 
quantity of spinal fluid is used than in the case of blood serum. It is our 
custom to use 3 pairs of tubes for each spinal fluid to be tested. The tubes 
in the front row contain antigen, complement and salt solution, and those in 
the rear row complement and salt solution only, just as in the tests on serum. 
To each pair of tubes are added respectively 0.2 c.c, 0.5 c.c. and 1.0 c.c. of 
the spinal fluid. The rear row of tubes serves as a control on any anti- 
complementary property which may be present in the fluid. In these 
control tubes, of course, the hemolysis should be complete. The results 
are noted and recorded exactly as described for patient's serum. 

When reading the results one first studies the controls. The corpuscles 
in the antigen-complement controls and in the complement controls should 
be completely hemolysed. The corpuscles in the tubes containing known 
negative serum should be completely hemolysed. The front tube of the 
pair containing a known positive serum should show no trace of hemolysis 
and its rear tube should show complete hemolysis. Any deviation from 
these results in the controls indicates that the results of none of the tests are 
dependable. In the case of the unknown sera a failure of hemolysis in the 
rear tube indicates that that patient's serum has properties which inhibit 
the action of complement, or is "anti-complementary." No result either 
positive or negative therefore can be recorded. If the corpuscles in the 
rear tube of a pair are completely hemolysed and there is no hemolysis or 
only partial hemolysis in the front tube, the result is read "positive." It 
is a common custom to read a result as + + + + if the corpuscles in the 
front tube show no trace of hemolysis ; to read + + + if there is very 
slight hemolysis; + -f- if approximately half hemolysis has occurred; and 
to read + if hemolysis is almost complete. 



582 



CLINICAL DIAGNOSIS 



The above is the procedure for testing a series of 16 sera with one antigen 
using the method for adjusting the reagents which we have found more 
delicate than many methods now in practice and which at the same time 
is not impractical since too time-consuming. If it is desired to use more 
than one antigen a third parallel series of tubes containing the second an- 
tigen, prepared exactly as described for the front row in the above series, 
is added to the "set-up." 

Interpretation. — It has been said that the strongest evidence of the value 
of the Wassermann test is the fact that now, but especially during the ear- 
lier period of its use, the results obtained by workers, many of whom pos- 
sessed little laboratory skill and less scientific training, were sufficiently con- 
sistent with the clinical diagnosis to establish the test in the position which 
it holds ; that is, the most valuable single laboratory test of the presence 
of syphilis. The question of the interpretation of the results obtained with 
the test has caused much debate among clinicians. Some hold, not without 
justification, that since the same serum tested separately in 2 or more 
laboratories may be reported with contradictory results, they cannot be 
expected to place reliance upon the test. This emphasizes the fact that 
before the question of its value can be discussed the qualifications of the 
serologist must first be considered. There are so many factors entering 
into the test and so many possibilities for error that unless the worker is 
thoroughly trained in the principles of serology and is conscientious and 
painstaking in his technic he may easily bring the test into disrepute. In 
discussing the interpretation of results, therefore, it is assumed that they 
were obtained by a method which is reliable and by a worker of unquestion- 
able accuracy. It is interesting that where such conditions obtain the 
number of reports of weakly positive reactions is reduced to a minimum 
and the most are definitely negative or strongly positive. Of course som 
sera will give a partially positive test even with most careful technic. The 
interpretation of these will be discussed later. 

Noguchi collected the results of a number of investigators who used the 
Wassermann reaction in the diagnosis of syphilis. The figures obtained by 
him are briefly summarized here since they still hold. 



Condition 

Primary syphilis 

Secondary syphilis manifest 

Tertiary syphilis manifest 

Early latent syphilis 

Late latent syphilis 

Hereditary syphilis 

Cerebrospinal syphilis 

General paralysis 

Tabes 



Number of 


Percentage 


cases 


positive 


416 


69.8 


1605 


89.4 


581 


78.I 


1233 


51. 


861 


47- 


125 


94-5 


64 


47.6 


498 


88.1 


2l6 


62.66 



THE BLOOD 583 

Noguchi reported a higher percentage of positive reactions in syphilis 
and the parasyphilides with his method of performing the test than with 
Wassermann's method. 

While the Wassermann reactions may be positive in a few conditions 
other than syphilis, this is not nearly as common as the earlier reports would 
indicate. The test is sometimes positive in frambesia (a disease caused by an 
organism very similar to that of syphilis), in malaria, in some cases of re- 
lapsing fever and in leprosy of the tuberculous type. Chloroform or ether 
anesthesia is occasionally followed by a positive Wassermann test in other- 
wise normal individuals. The early statement that the blood in scarlet 
fever sometimes gives a positive test has not been substantiated, for now 
it is found uniformly negative except in cases where syphilis cannot be 
ruled out; and the same is true of pellagra. 

Malaria with Positive Wassermann. — J. S., No. 6858, aged 13 years, was admitted 
with a diagnosis of tuberculous coxitis (left). The high fever, sometimes accompanied 
by a chill, suggested malaria and blood examination demonstrated Plasmodium vivax. 
On Sept. 21, 19 1 8, the Wassermann test of the blood was 3 plus and Sept. 28th it was 
4 plus. Quinine was started September 22nd. On both Oct. 5th and 12th the blood 
Wassermann was negative. 

Treatment with mercury or salvarsan may quickly cause the serum 
of a syphilitic patient to lose the power of giving a positive Wassermann 
reaction, so that for diagnostic purposes it is necessary to take this fact 
into consideration. 

In cases suspected of cerebrospinal lues including tabes and paresis it 
is advisable to test both the serum and spinal fluid , since either one and not 
the other may prove positive. 

With the exceptions of the conditions noted above the present opinion is 
that a positive Wassermann test indicates the presence of living spirochetes 
somewhere in the tissues of the patient. 

Unfortunately a negative reaction cannot be interpreted as definitely 
as can one which is positive. Even with the most painstaking technic and 
using every modification of the test the reaction in a small percentage of 
known cases of syphilis in all stages is negative. It has been demonstrated 
repeatedly that an alcoholic debauch will occasionally produce a negative 
reaction in an individual whose serum previously gave a positive reaction. 
The percentage of negative tests varies much in different stages of the 
disease as well as in the reports of different workers. The percentage of 
these erroneous negative reports has become progressively smaller as the 
technic cf the test has improved but there is at present no reason to hope it 
will be reduced to zero. 

It has been shown that the amount of the substance in the syphilitic 
patient's serum which absorbs complement in the presence of appropriate 
antigen varies widely. For illustration, in some cases of unquestionable 
luetic infection no such substance can be demonstrated, while in other simi- 



584 CLINICAL DIAGNOSIS 

lar cases it is present in amounts sufficient to. absorb several times the 
quantity of complement ordinarily used in the test. It is easy to see that 
between these two extremes may be found all possible quantitative vari- 
ations in the complement-absorbing power of the serum of a syphilitic 
patient. It therefore would not be strange if in certain cases of syphilis 
the serum contained enough of this substance to bind all the complement 
used in the test and in addition left enough free to cause partial hemolysis 
when the indicator is added. -If the presence of this substance in quantity 
sufficient to absorb all the complement, and therefore give a -f- + + -f- 
positive reaction, is evidence of syphilis, then the presence of this sub- 
stance in any detectable quantity whatever is similar evidence. Therefore, 
if the technic is above question, a-f+ora + -f + reaction should be re- 
garded as of practically the same significance as a + + -f- -f- positive. A 
faint positive, as + , should, however, be interpreted as suggestive only. 

Complement Fixation in the Diagnosis of Gonococcus Infection. — Very- 
early in the development of the complement fixation test attempts were 
made to apply it to the diagnosis of gonococcal infection but these proved 
only partially successful. Later it was found that the gonococcus is not a 
single distinct strain or species, but a large group of closely allied strains of 
organisms which can be differentiated by the agglutination tests. The 
antigens used in the earlier experiments presumably were prepared from 
only one of these strains. Later investigators found that by using antigen 
containing each of the known strains of gonococcus — i. e., polyvalent 
antigens — the complement fixation test gives satisfactory results. 

The difficulty of isolating the gonococcus in pure culture and the care re- 
quired in sub-culturing a large number of strains make the preparation of 
gonococcus antigen so tedious that most workers use an antigen prepared 
and standardized by a laboratory whose specialty is the preparation of 
biological products. 

As a preliminary step the amount of the antigen necessary to show anti- 
complementary properties is carefully determined, using the same scheme 
as described for testing Wassermann antigens, and a quantity not larger 
than % this amount is used in the test. Complement, amboceptor and 
sheep corpuscles are prepared and standardized exactly as for the Wasser- 
mann test. It is important that whatever hemolytic system is used it 
should be so adjusted that the amount of complement should be small 
in proportion to the amount of patient's serum used. For this reason, 
as in the Wassermann test, one-fifth the original Wassermann quantities 
are to be recommended. The patient's serum is inactivated as for the 
Wassermann test. 

One pair of tubes is allowed for each patient's serum and i pair for 
each control. One-tenth of a cubic centimeter of complement so diluted 
that this amount contains 2 units, is measured into every tube in the 



THE BLOOD 585 

series. One-tenth of a cubic centimeter of antigen, so diluted that this 
amount contains not more than % the anti-complementary dose, is added 
to each tube in the front row. One-tenth of a cubic centimeter of each 
patient's serum is placed in each of the pair of tubes assigned to that test. 
To t pair of tubes is added a previously tested positive serum and into 
another pair a negative serum. An additional pair contains no serum, but 
the front tube antigen and complement in the above specified quantities, 
and the rear tube the complement alone. The total volume in each tube is 
brought to 0.5 c.c. by the addition of salt solution. The set-up is incubated 
in a water bath at 3 8° C. for 1 hour, after which 0.5 c.c. of a 2% suspension 
of sheep corpuscles and 2 units of the proper amboceptor are added to each 
tube. The incubation is repeated and the results recorded exactly as de- 
scribed for the Wassermann test. 

The complement fixation test for gonococcus is a definitely specific test 
which depends on the presence of specific antibodies in the patient's serum. 
It therefore differs from the test for syphilis. The test is frequently negative 
in acute gonococcal urethritis, probably because the infection, being acute 
and local, has stimulated the production of less free antibody in the cir- 
culating blood than would one which has extended to other structures and 
which has been of longer duration. The highest percentage of positive 
results is obtained in series of cases of acute exacerbations of chronic ure- 
thritis, in involvement of the prostate or epididymis, in gonorrheal iritis, in 
salpingitis and in arthritis. In these conditions the serum of approximately 
80% of the cases gives a positive test. A failure of the patient's serum to 
fix complement in the presence of gonococcus antigen, therefore, does not 
exclude gonococcus infection. A positive reaction may persist for several 
weeks after all evidence of the infection has disappeared. 

Williams 109 found the test of particular value in strengthening one's sus- 
picion of a gonococcal infection in cases in which the gonococcus could not 
be discovered bacteriologically, nevertheless the final proof of a gonorrheal 
infection is the cultivation of the gonococcus. As a matter of technic the 
cultivation of the gonococcus, owing to its susceptibility to cooling, its 
association with other more rapidly growing organisms and the need of 
special media, is much more difficult than the fixation test. Except in acute 
cases, where the gonococci are abundant, the examination of a Gram-stained 
smear should not be accepted as conclusive since Micrococcus catarrhalis, 
Diplococcus crassus, irregular types of Gram-negative cocci, and, most 
important of all, the so-called " degeneration" forms of staphylococci may 
lead to error. 

In interpreting a positive test, it should always be borne in mind that 
gonorrhea is a very widespread disease and that an individual may suffer 
from at least two different infections. 

109 Interstate Med. Jour., xxi, 19 14. 



586 CLINICAL DIAGNOSIS 

A positive reaction in a patient supposedly cured of gonorrhea indicates 
the presence of a gonococcal focus and his capability of infecting others. 
The importance of this in connection with problems of marriage is great. 
A positive reaction occurs in about 20% of those clinically cured. 

In acute cases, in which the gonococci are usually easily demonstrated, 
the fixation test is generally negative. On the contrary in the chronic and 
ill-defined affections, where it is not often possible to obtain the organism, 
the test acquires its greatest sensitiveness, expecially in -the diagnosis of 
gonorrheal arthritis and of gonorrheal epididymitis, at least by the fifth 
week, about 100% of which cases give positive reactions. About 75% of 
the cases of posterior urethritis, prostatitis and seminal vesiculitis with re- 
current exacerbations, about 65% of cases of pyosalpingitis and about 65% 
of all stricture cases give positive reactions. 

A negative reaction would not exclude gonococcal infection, especially 
in the acute and subacute stages without complications and when limited 
to the urethra or vagina. 

Syphilis or a positive Wassermann reaction does not interfere with 
the test. 

In gynecology the test has proved its value in the differential diagnosis 
of pelvic inflammatory diseases from one another and from neoplasms. 
The test is usually negative in uncomplicated cases of urethritis, vulvo- 
vaginitis and Bartholinitis ; it appears that the infection must reach at least 
to the level of the uterus before a positive reaction develops. 

Complement Fixation for Tuberculosis. — Attempts had been made to 
demonstrate complement fixation in cases of tuberculosis before Wasser- 
mann applied the test to syphilis. In these earlier attempts Koch's tuber- 
culin and other similar preparations were used as antigen. Subsequently 
a great variety of preparations of tubercle bacilli and their products were 
• used by various workers and with various results. The literature on this 
subject is too extensive to be reviewed here, but in brief it may be said that 
antigens prepared by widely different methods have given a fairly high 
percentage of positive tests in cases of tuberculosis, especially the early 
cases, and that the percentage of positives in normal individuals is small. 
Our own experience with the test has led us to believe that if extreme care 
is used, especially in adjusting the hemolytic system, the test is of real 
value in diagnosis. It should be regarded in the same manner as other 
laboratory tests, not as evidence on which alone the diagnosis of the pa- 
tient's present illness may be established, but as evidence which, properly 
interpreted, will help. We have used antigens prepared after the methods of 
Miller, Bronfenbrenner and Petroff in a large series of cases and nave found 
the results to conform closely with the results of the clinical examination. 

The wide variation in the results obtained by various workers with this 
test is probably due to the fact that in tuberculosis free antibodies are 



THE BLOOD 587 

frequently present in the blood in amounts too small to bind the relatively 
large amounts of complement which some use in complement fixation 
technic. For example, many use in the hemolytic system 0.5 c.c. of a 
5% suspension of sheep corpuscles. Two of the units of complement which 
would correspond to this quantity of sheep cells would be a larger amount 
than could be absorbed by the amount of tuberculous antibodies usually 
present unless these happen to be present in unusually large amounts. If, 
on the other hand, the hemolytic system is based upon 0.5 c.c. of a 2% 
corpuscle suspension, two units of complement standardized for this 
quantity might be absorbed by the patient's serum in the presence of suit- 
able antigen and result in a positive test. The above explanation has been 
justified repeatedly both in tests for syphilis and for tuberculosis by finding 
sera which react negatively or faintly positively with the larger units of 
complement but which completely bind the complement when a smaller 
unit is used. 

The simplest method of preparing an antigen for the test for tubercu- 
losis is that of Miller. A number of strains of tubercle bacilli are grown in 
glycerine broth, and the growths from the cultures are collected and mixed. 
Of this, 10 mgms. is mixed with 90 mgms. of dry sodium chloride and ground 
for an hour in an agate mortar. This amount is made up to isotonicity with 
distilled water and vigorously shaken. The supernatant fluid after the 
larger particles have settled to the bottom of the tube is the antigen. It 
is well for the sake of safety to sterilize this by heat before using it. The 
test for the anti-complementary property of the antigen, the preparation 
of reagents and the technic of the test are exactly the same as described for 
the Wassermann test. 

It has been found that a considerable percentage of the serums which 
give a strongly positive Wassermann test will bind complement also with 
tuberculous antigens. This is spoken of as "cross fixation" and makes it 
necessary when applying the test for tuberculosis to test each serum first 
against a syphilitic antigen, otherwise a cross fixation might be mistaken 
for a positive tuberculous fixation. Stivelman n0 from the study of 700 
cases considered the test of little clinical value since but 33% of the cases 
of definite incipient tuberculosis gave a positive test and since it did not 
assist in determining clinical activity or immediate prognosis. Petroff m 
believes this test even more specific than the Wassermann. 

It should be pointed out that there is no necessary conflict in these 2 opinions. The 
test may be very specific and yet of little value. We take it for granted that a much 
larger percentage of persons have at some time of their lives had tuberculosis of which 
they were not aware and of which they now are well than is the percentage of those who 
have known of their infection because of outspoken symptoms. The question is, Have 
they now a tuberculous infection which they should fight? 

110 The Jour, of Lab. and Clin. Med., April, 1920, v, p. 453. 

111 Am. Rev. of Tuberc, 1920, iii, p. 683. 



588 CLINICAL DIAGNOSIS 

ISOHEMAGGLUTININS 

The presence of isohemagglutinins in human blood was discovered 
independently by Landsteiner and Shattuck in 1900. Since then ex- 
tensive studies of these have been made by a number of workers. Land- 
steiner found that individuals may be grouped into 3 classes according to 
the behavior of their serums and corpuscles toward those of other in- 
dividuals. A fourth group or class has been added by subsequent investi- 
gations. In 19 10 Moss published an extensive study of isohemagglutinins. 
He divided individuals into 4 groups according to the behavior of their 
bloods and determined the percentages of individuals in the different 
groups. Unfortunately Moss and Landsteiner, whose results are practically 
identical, named their groups differently, Moss assigning to group four 
the individuals placed by Landsteiner under group one. Confusion in- 
evitably arose from this situation. 

Landsteiner's classification is as follows: 

Group I. — The corpuscles of Group I are not agglutinated by any human 
serum; the serums of Group I agglutinate the corpuscles of each of the 
other groups. 

Group II.— -The corpuscles of Group II are agglutinated by the serums 
of Groups I and III; the serums of Group II agglutinate the corpuscles of 
Groups III and IV. 

Group III. — The corpuscles of Group III are agglutinated by the serums 
of I and II; the serums of Group III agglutinate the corpuscles of Groups 
II and IV. 

Group IV. — The corpuscles of Group IV are agglutinated by the serums 
of each of the other groups ; the serums of Group IV do not agglutinate the 
corpuscles of any other groups. 

According to , the classification of Moss the groups are arranged 
as follows: 

Group I. — The corpuscles of Group I are agglutinated by the serums 
of Groups II, III and IV; the serum of Group I does not agglutinate any 
corpuscles. 

Group II. — The corpuscles of Group II are agglutinated by the serums 
of Groups III and IV; the serum agglutinates the corpuscles of Groups 

I and III. 

Group III. — The corpuscles of Group III are agglutinated by the serums 
of Groups II and IV; the serum of Group III agglutinates the corpuscles of 
Groups I and II. 

Group IV. — The corpuscles of Group IV are not agglutinated by any 
serums. The serum of Group IV agglutinates the corpuscles of Groups I, 

II and III. 

These relationships may be graphically demonstrated by the accom- 
panying charts: 



THE BLOOD 

Landsteiner 



589 





Series I 


Series II 


Series III 


Series IV 


Corpuscle I 











O 






Corpuscle II 


+ 





+ 





Corpuscle III 


+ 


+ 


o 


o 


Corpuscle IV 


+ 


+ 


+ 


o 






Moss 


Corpuscle I 





+ 


+ 


+ 


Corpuscle II 


o 


o 


+ 


+ 


Corpuscle III 





+ 





+ 


Corpuscle IV 





o 









+ = Agglutination. 
o=No agglutination. 

The following approximate percentages are given to the different groups 
by Moss: Group I, io%; Group II, 40%; Group III, 7%; and Group IV, 
43%. It is stated that the type of blood of the individual is an inherited 
trait which is transmitted according to the Mendelian law and is permanent 
during the life of the individual. 

The use of blood transfusion as a therapeutic measure has brought 
the subject of hemagglutinins into prominence since it is found that 
frequently serious results follow when donor and recipient are members 
of different groups. 

It was formerly the custom to test the bloods of donor and of recipient 
for compatibility by mixing the donor's corpuscles with the recipient's serum 
and the donor's serum with the recipient's corpuscles and examining these 
mixtures for the presence of agglutination. If no agglutination appeared 
the bloods were pronounced compatible. 

The above procedure, while perfectly satisfactory, has been superseded 
in large measure by the test for the determination of the group to which an 
individual belongs. To do this it is necessary to have serums known to 
belong to Groups II and III. Approximately 2 drops of the blood of the 
individual to be tested are suspended in about 1 c.c. of salt solution which 
contains preferably about 0.5% of sodium citrate. This gives a corpuscle 
suspension of suitable density. Upon a clean cover glass are placed a 
loopful of serum Type II and a loopful of serum Type III. A loopful of 
the corpuscle suspension is added to each and a third loopful of the cor- 
puscle suspension is placed on the same cover slip as a control. A hollow 
ground slide rimmed with vaseline is inverted over the drops on the 



590 CLINICAL DIAGNOSIS 

cover slip and after 5 to 10 minutes the preparation is examiner', under low 
power of the microscope. The presence of agglutination is easily detected 
by inspecting the corpuscles. It will be seen by reference to the above 
charts that the group of any individual's blood may be determined by the 
behavior of its corpuscles to the serums of Groups II and III. Thus, in 
Moss's classification, if the corpuscles are agglutinated by both serums II 
and III, the blood belongs to Group I ; if the corpuscles are agglutinated by 
serum III and not by serum II, the blood belongs to Group II; if the 
corpuscles are agglutinated by serum II, and not III, it belongs to Group 
III ; and if the corpuscles are agglutinated by neither serum it belongs 
to Group IV. 

It is seen that the determination of the group to which donor and re- 
cipient belong is a simpler procedure than that of testing them for hemag- 
glutination by crossing the serum of each with the corpuscles of the other. 
It has the additional advantage also that the type of blood of the recipient 
need be tested but once. Each prospective donor is then tested sepa- 
rately until one is found belonging to the same group as the recipient. 
The sera used for the agglutination test will keep for many weeks 
without deterioration. 

THE BLOOD IN DISEASE 

Anemia. — The popular definition of an anemia has been, a deterioration 
of the blood qualitatively and quantitatively as regards one or all of its con- 
stituents — the plasma, the corpuscles and the hemoglobin (Grawitz) . While 
the red blood-cells are in fact a relatively unimportant part of the total 
blood compared with the plasma upon which depends the health of the 
whole body including the red corpuscles, yet their number and their hemo- 
globin are the only easy criteria for estimating the blood's condition and so 
practically the term anemia is limited to conditions of the blood which 
affect these cells, their number per cubic millimeter, their hemoglobin con- 
tent, or both. Unfortunately these values relate to but 1 c.mm. of blood 
and not to the volume of blood as a whole. 

The concept of anemia should include a diminution in the total volume 
of the blood but the estimate of this is as yet not practicable clinically 
although examinations at the autopsy table even in cases with a practically 
normal blood-count show that this volume does change considerably. 

Many have tried to define anemia in terms of changes in the plasma 
but this has proved unsatisfactory. Some definite plasma changes may 
mean little while the plasma in the severest anemias (clinically) with the 
lowest counts may chemically be almost normal. 

By oligocythemia or hypocythemia is meant diminution in the number of 
red blood-cells in 1 c.mm. of blood. This may be due either to an actual 
reduction in the total number of the red cells in the body, or merely to an 
increased volume of plasma. By oligochromemia is meant a diminution in 
the amount of hemoglobin as judged by its color compared with that of an 



THE BLOOD 591 

equal volume of normal blood. By color -index is meant the percentage of 
hemoglobin divided by the percentage of the red blood-cells, 5,000,000 
cells considered as 100% (see page 482). 

By oligemia is meant a diminished total amount of blood in the body. 
This may be suspected, but as yet cannot be proved. Oligemia serosa is an 
oligemia of diluted blood; oligemia sicca, an oligemia with blood qualitatively 
normal. Hydremia means an increased percentage of water in the plasma 
and occurs whenever the albumin is diminished. Polyplasmia is an increase 
in the volume of the plasma, supposed to occur in chlorosis; oligoplasma, 
a decrease, which occurs in certain cardiac diseases. By plethora vera is 
meant an increase in the total volume of blood. This can only be suspected. 

By the hematopoietic organs one usually means the organs furnishing the 
corpuscles; i. e., the bone-marrow, spleen and lymph-glands. The bone- 
marrow certainly furnishes red corpuscles and many leucocytes ; the spleen 
is active perhaps after a severe hemorrhage, but otherwise is probably un- 
important in the production of red blood-cells. It is possible that some 
leucocytes originate there and quite probable that this organ removes some 
of the old cells. The function of the lymph-glands as hematopoietic organs 
is still in doubt. 

The red blood-cells have the indispensable yet simple function of trans- 
porting the oxygen to the tissues. The leucocytes are thought to be 
important in immunity production and in the absorption of neutral fat 
in the intestine, while by their disintegration they certainly raise the al- 
bumin content of the blood. The function of the platelets, apart from their 
possible relation to coagulation, is not understood. While these func- 
tions of the formed elements are very important those of the plasma of the 
blood are far more intricate and varied . We study the former because they 
are as yet the only practical index we have of the condition of the latter. 
Yet in studying the anemias clinically the organs which form the plasma 
must be most considered ; especially the intestine, the liver and the kidneys. 
It is in the intestinal wall that the plasma obtains its proteid content; in 
the liver that many processes are brought about including the control of the 
carbohydrate content of the plasma, the transformation of the ashes of the 
body to urea, etc. ; and it is in the kidneys that the most of the ashes are 
removed. Certain of the glands of internal secretions also are important in 
modifying the constitution of the blood, especially the pancreas, thyroid and 
the adrenal. Lastly, in the muscles themselves the blood is modified since 
they remove certain tissue constituents and glucose and give back in return 
the ashes of these bodies. 

In many diseases of the blood it" is probably the plasma that suffers 
first, while the changes in the red blood-cells are the results of the plasma 
changes but also of secondary changes in the blood -building organs. 

The anemias have been classified as primary and secondary. By pri- 
mary anemia was meant one which seems to develop independently of any 



592 CLINICAL DIAGNOSIS 

organic disease, e.g., chlorosis, the essential idiopathic anemias (the simple 
primary and the pernicious anemia, leukemia and pseudoleukemia). By 
secondary anemia was meant one for which an adequate cause seems pres- 
ent, as hemorrhage, blood poisons and organic diseases as lues or cancer. 
The above classification is purely clinical, not hematological, and the au- 
topsy table not infrequently shows to be secondary an anemia which was 
during life supposed to be primary. Lately the terms primary and second- 
ary relate more to the blood pictures or types rather than to the cause 
and are being replaced by the terms megaloblastic, chlorotic, etc. 

By hypoplastic or aplastic anemia is meant one due to insufficient 
blood formation; by consumptive or hemolytic anemia, one due in part 
at least to increased blood destruction. 

Secondary Anemia. — A secondary anemia is, from the point of view of 
the pathologist, one which can be assigned to some cause which would seem 
adequate to explain the condition. But to the hematologist the term suggests 
a blood the most of the red cells of which are smaller and of lighter weight 
than normal, while a few are large, pale and "waterlogged." We usually 
may make out certain features which have in the past suggested that a 
toxin has injured the red cells: basophilic granular degeneration, poly- 
chromatophilia, etc., but these may just as well be evidence that these cells 
are immature. The light-weight cells would suggest that the formation of 
new hemoglobin is a much more difficult function than is the multiplication 
of new cells (granting that the material is at hand out of which these new 
cells may be made). In response therefore to an unusual demand on the 
erythroblastic tissue for new cells the hemoglobin available would seem to be 
divided into a large number of light-weight cells rather than into a smaller 
number of normal cells. This we can understand since from the point of 
view of the physiologist the area of surface of the cells is of more importance 
than is their mass. As the patient improves the light-weight cells are grad- 
ually replaced by those of normal weight, therefore the color index ap- 
proaches i. 

In mild cases of secondary anemia the count may be normal but the 
hemoglobin is diminished and the specific gravity slightly lowered since 
many of the cells are light-weight, i. e., are small and pale. In cases a little 
more marked (moderate grade) the count may be normal, but the reds are 
not only light-weight but show qualitative changes: degenerations (?), 
anisocytosis, poikilocytosis, crenation, poly chromatophilia and less tendency 
to rouleaux formation. In severe cases the blood shows both qualitative 
and quantitative changes but the count is not much reduced except in the 
anemias of childhood, after large hemorrhages, in malaria and in acute sep- 
ticemia. In very severe cases one sees also the evidences of degeneration (?) 
and destruction (?) of the cells and signs of regeneration (nucleated reds). 

Blood Picture. — In secondary anemia the blood may grossly be pale. 
The reds are less reduced than is the hemoglobin ; their count may even be 



THE BLOOD 593 

normal, but in severe cases there will be a great reduction, as In v. Limbeck's 
case with recovery with a count of 306,000. A reduction of 1 ,000,000 cells is, 
Bezancon and Labbe consider, a mild hypocythemia, 1 of from 2,000,000 
to 3,000,000 an intense; while if the cells are reduced to 1,000,000 an ex- 
treme hypocythemia. 

1 The reduction in hemoglobin is the constant and most important fea- 
ture and the best index of the grade (yet see page 503) of a secondary 
anemia. The color-index is low in cases due to cancer, hemorrhage and 
gangrenous processes, yet not quite so low as it is in chlorosis. On the other 
hand in cases with extreme oligocythemia the body, if given sufficient time, 
seems to protect itself by increasing the color-index, that is, by the pro- 
duction of cells which in size or weight are normal or above normal. Some 
think that the high color-index of pernicious anemia itself is not character- 
istic of the disease, but a reaction to the low count ; the body, because of the 
chronicity of the disease, having had time to thus protect itself, while in 
those cases of secondary anemia with low count and low colcr-index the 
acute course prevents this protective measure. Another suggestion we 
would make is that because of abnormal hematopoiesis the bone-marrow, 
unable to use in new cells all the pigment liberated by the normal breaking 
down of worn-out cells, saves some of this hemoglobin by overloading the 
cells it can produce. The specific gravity of the blood is low. The dried 
residue is reduced. This is especially true in the cancer cases. (In 1 case of 
cancer of the stomach with a count of 1,400,000 and 15% Hb the dried 
residue was only 9%.) And yet a lowering of the red cell-count and of 
the hemoglobin may be a sign of improvement. This is well seen in some 
cases of anemia with a reduction of the total volume of the blood. During 
the convalescence in such cases the blood volume is first restored by an 
increase of plasma which dilutes the blood and gives the appearance of a 
progressing anemia. 

Morphologically the stained cells show a lack of hemoglobin and yet a 
good many are normal. In many the biconcavity is too evident and pessary 
forms are common. The polychromatophilic degeneration is common and 
is seen within 24 hours after a hemorrhage, but bears no relation to the 
hemoglobin-content of the cell. The number 01 these basophilic cells 
runs so parallel to the grade of the anemia that the estimation of their 
number has been suggested as a substitute for the more difficult blood- 
counting (Walker). 

Poikilocytes occur only in the severest cases. Anisocytosis is marked. 
Microcytes are always found, some even but 2 11 in diameter. Large cells 
"acutely dropsical" have been described. 

The number of nucleated reds varies much and bears no relation to the 
anemia, either to its grade or its cause. They may be abundant or absent. 
Blood crises, normoblastic as a rule, are not rare (see page 606). Micro- 
blasts are met with in the severe post-hemorrhagic type and megaloblasts 
38 



594 CLINICAL DIAGNOSIS 

are exceedingly rare except in cases due to malaria and other diseases which 
affect the bone-marrow. 

The number of the leucocytes varies from a leucopenia to a leukemic 
condition, depending on the cause of the anemia and its complications. 
Duiing convalescence a moderate leucocytosis with an increase in the 
polymorphonuclear neutrophiles often develops, due to the increased ac- 
tivity of the bone-marrow. The number of eosinophiles varies much, from 
few to an extreme eosinophilia. As a rule their count is at the upper limits 
of normal. 

The platelets are increased, even doubled in number. This is always 
true in the post-hemmorrhagic cases. 

Acute Post-Hemorrhagic Anemia. — An anemia due to hemorrhage 
may be acute or chronic depending on the number of hemorrhages 
and on the length of the intervals between them. The loss at one time 
of ){ to % the volume of blood is fatal. Women tolerate hemorrhages 
better than men and children least well of all. 

Immediately after a hemorrhage the blood is for a while qualitatively 
normal; then, as the tissue-lymph pours into the vessels to restore the 
blood volume, the count and the hemoglobin diminish and the specific 
gravity becomes somewhat lower since tissue lymph is richer in water 
than is the plasma. The loss of even but 50 to 70 c.c. of blood is followed 
by demonstrable changes. The color index therefore should for a short 
time remain "1" then will decrease, since the new cells are "light weight," 
smaller in size, paler in color than normal and abnormal both in shape 
and staining qualities. The red cell-count then rises slowly until normal 
and the hemoglobin more slowly since it is weeks before the light-weight 
cells can be replaced by new ones of normal volume. 

The platelets are increased in number. After 1 hemorrhage the hy- 
dremia is maximal and the color-index minimal on about the ninth day. 
There is often a post-hemorrhagic leucocytosis. The regeneration of the 
red blood-cells is so rapid at first that some suspect direct division of some 
of the red blood-cells of the circulation and point out in favor of this the 
number of small cells and of poikilocytes which appear so early. 

One usually finds a few nucleated reds, usually normoblasts, their num- 
ber related more to the acuteness of the hemorrhage than to its severity, 
while during convalescence blood crises are common (see page 606). 

In 1 case of severe post-hemorrhagic anemia 13.7% of myelocytes were 
found in the circulation. These disappeared in 3 days. In another very 
severe case the polymorphonuclears were free from granules. 

An early feature in the regeneration of red cells is the production of so 
many megalocytes that in some cases they are a conspicuous element oi 
the blood-picture. 

Time for Regeneration After One Hemorrhage. — The table given by v. 
Limbeck is : 



THE BLOOD 595 

If the loss of blood was 4.5% of the body weight 30 days are necessary 
for complete regeneration; if 4% of body weight, 20 days; if 3%, 10 days; 
and if 2%, 8 days. 

Grawitz says a loss of 3 to 4% of body weight requires from 14 to 30 
days for regeneration; one of 1 to 3% from 5 to 14 days; a slight loss from 2 
to 5 days. 

But the time varies also with the age and nutritional condition of the 
patient, the diet and the therapeutic measures used. 

Regeneration is quickest in men between 20 and 40 years of age; slower 
in women and slowest in children. After the regeneration is complete there 
may develop even a hypercythemia. 

In 1 case due to repeated hemorrhages following abortion (produced evidently by- 
some drug, not by an operation) the count on admission (the temperature was normal) 
was 1,108,000, hemoglobin 18% and the leucocytes 4625. 

In a case of hemorrhage from a badly crushed arm and after infusion the red cells 
fell during 36 hours from 5,000,000 to 3,000,000, and the hemoglobin from 70 to 50%. 
The hemoglobin in a case of metrorrhagia fell to 19,% and in another case after 2 post- 
partum hemorrhages to 11%. Both patients recovered. 

Among the causes of acute anemia are: traumatic hemorrhage, tubal 
pregnancy (in which a rapid anemia is a bad sign), abortion, uterine sub- 
mucous tumors, ulcers of duodenum and stomach, typhoid ulcers, phthisis, 
aneurisms, varicose veins of esophagus, rectum, or legs, the ''hemorrhagic 
diatheses " and hemorrhagic pancreatitis. 

A case of purpura hemorrhagica of eight weeks' duration 112 was admitted 
with red cells 696,000, hemoglobin 17% and leucocytes 4000 (s. m. 75%). 
At death 7 days later the red count was 483,000, there were no poikilocytosis 
(since too acute?), no nucleated reds and no eosinophiles. 

Ewing mentions a case of 3 weeks' duration with repeated epistaxis and 
a red count of 456,000. 

In this group of the acute hemorrhagic anemias are included cases of 
r2peated hemorrhages, yet with hemorrhages at such intervals that com- 
plete regeneration after each is possible. In these cases the total amount of 
blood lost may be enormous. This was well seen in the days when venesec- 
tion was a common practice. Ehrlich mentions a Russian physician 
with pulmonary tuberculosis who, in 6}4 months, lost 20 kilos of blood 
(that is, an amount 4 times the total blood volume at any time) and yet 
recovered perfectly. 

Anemia from Chronic Hemorrhage. — By chronic hemorrhage is 
meant a succession of hemorrhages at such intervals that the patient can- 
not recover from one before the next occurs. The results are much more 
serious than those following 1 hemorrhage, or a series with longer intervals, 
although the total amount of blood lost may be relatively small. A case 
with repeated epistaxis due to telangiectasis of the nasal mucosa was ad- 

112 Billings, Johns Hopkins Hosp. Bull., May, 1894. 



596 CLINICAL DIAGNOSIS 

mitted several times, once with red cells 2,288,000, hemoglobin 18% and 
leucocytes 2800. Scurvy, especially cases with much hemorrhage, causes a 
secondary anemia, the red count averaging from 3,000,000 to 4,000,000. 
In 1 severe case it was reported as low as 370,000. A leucocytosis is often 
met with in the chronic post-hemorrhagic anemias but is due to some com- 
plication, since a leucopenia is the rule. In one case in this clinic the red 
cells were 2,200,000, hemoglobin 40% and leucocytes 2850. This form of 
anemia is common in cases of pulmonary tuberculosis, submucous fibro- 
mata, hemorrhoids, gastric and intestinal ulcers, cancers (e. g., of the stom- 
ach), intestinal parasites, etc. The severe anemias from high and hidden 
piles is now attracting much attention. 

As a result of a long-standing anemia the blood-building organs seem to 
lose their ability to regenerate the blood and convalescence is therefore very 
slow. In 1 case of hemorrhoids with a count of 2,600,000 it required 8 
months to reach normal (Ehrlich) . Sometimes a chronic secondary anemia 
assumes the picture of a primary anemia, sometimes without any signs of 
regeneration and is rapidly fatal. That these anemias are due in part to the 
chronic diseases which cause the hemorrhages is one reason why it takes 
much longer for the blood in these cases to regenerate. 

In this form of anemia the hydremia is marked, the specific gravity of 
the blood is low and the dried residue considerably diminished. The red 
count is much diminished, even to 1,000,000 cells. The new reds are small 
and pale and the index low, 0.5 or even 0.44. Nucleated reds are scanty 
and the platelets are increased. Sometimes the picture becomes that of a 
pernicious anemia (it may be, that other patients die before this blood pic- 
ture can develop) , yet in fatal cases the index usually falls progressively 
until death. The leucocytes often are increased at first but when the 
anemia becomes very severe a leucopenia usually exists. 

Blood Poisons. — Certain poisons may cause anemia by destroying the 
corpuscles themselves or by injuring the blood-building tissues. Among 
them are the toxins of the infectious diseases, especially the septicemias, 
scarlet fever and lues; certain mineral poisons, as lead, arsenic and mercury; 
the toxins of intestinal parasites, as Bothriocephalus latus; and especially 
the toxin of malignant tumors. 

The effect of these poisons is sometimes seen in various degenerations ( ?) 
of the red blood -cells in the circulation and in the hemoglobinemia (plas- 
molysis) . Other toxins are thought not to injure the cells in the circulation 
but to cause an increased activity on the part of the blood-destroying organs, 
the liver, the spleen and the marrow. The best illustration of the effects 
of such a hypothetical toxin is hematochromatosis. It may be that the 
poison produces a chemical (plasmotropic) change in the protoplasm of the 
red cells which singles them out for destruction. 

Anemia op Inanition; the Anemia of the Poor. — The anemia seen 
so often among the poor is considered by some as a simple primary anemia, 



THE BLOOD 597 

by others as a secondary anemia due to a variety of concurring factors the 
relative importance of which cannot be apportioned, such as poor food, 
lack of sunlight, bad air, worry and overwork. The present idea is that it 
is due to long-standing latent infection. 

Starvation alone will not cause an anemia ; that is, it will not produce 
qualitative changes in the blood but causes rather a diminution in the total 
volume of blood which runs roughly parallel to the loss of weight. The 
blood of Cetti, at the end of a 10-day fast, showed a rise in the red blood-cell 
count of 1,000,000, a slight fall in the hemoglobin, while the leucocytes 
fell from 1 2 ,000 to 4200. In such cases the blood picture of anemia may be- 
gin with the improvement of the patient's condition after he begins to 
eat since the formation of new blood is a slower process than the gain of the 
other tissues. The first blood changes as these patients improve will be 
those of dilution, i. e., an apparent anemia. 

Poor food has been considered an important cause of chronic anemia of 
the hypoplastic form (Immermann) , i. e. , of anemia due to insufficient blood 
formation. In support of this view it is emphasized that the foods which 
contain most iron are the most expensive and that anemia is met with 
particularly in those European countries where the diet of the poor con- 
sists chiefly of bread, potatoes, etc. And yet this cause is greatly exag- 
gerated. In this country at least the trouble is not so much the quality of 
the food, since even the poorest contain enough of this metal to replace 
that which daily is actually lost to the body, as its preparation, good meat 
and vegetables being rendered indigestible in the frying-pan; and also the 
American habit of eating in a hurry and insufficiently masticating the food, 
all of which would cause gastro-intestinal troubles which might result in 
anemia. Bunge's experiments showed that a diet poor in iron can cause 
anemia in a growing child, but the problem of growth is a different one. 
The effect on the blood of a non-proteid diet, however, can be demon- 
strated at the end of 6 or 8 days, at first by a slight hydremia and later by 
the changes in the red blood-cells characteristic of a secondary anemia. 

Those living in dark rooms are very apt to be anemic, not because of the 
lack of sunlight but because of the prevalence of tuberculosis, an anemia- 
producing disease common among those seldom in the sunshine. Hemo- 
globin is not comparable to chlorophyll as the illustrations cited by Ehrlich 
show. The horses which were kept at the bottom of mines in Germany for 
from 10 to 24 years without seeing sunlight had normal blood; the members 
of Nansen's Polar Expedition remained for 140 to 150 days without sun- 
light and yet remained healthy provided the other causes of anemia were 
eliminated. On the other hand although sunlight may not be very important 
for the adult it is for the growing organism (Schonenberger) , but other 
factors also are potent to explain this. 

Life in an atmosphere of bad air also would seem to predispose one to 
anemia, while overwork and worry would superficially, at least, seem im- 



598 CLINICAL DIAGNOSIS 

portant causes of the anemia of the poor. Many so-called ' ' scientific facts " 
which have been useful in the propaganda of a reform are held tenaciously 
by reformers who will continue for a long time probably to assign to the 
above factors the anemia of the poor. And yet indirectly these are its causes 
and their correction would do much to eliminate this form of anemia and 
also the anemia of the rich since they lower the resistance of the patients to 
the various latent infections of lungs, kidneys, teeth, tonsils and bowel 
and it is these which cause anemia. 

There is a group of simple secondary anemia for which no one cause can 
be assigned. The great majority are met with in women. In one series the 
mean of the red cell-count was about 3,000,000 (2,100,000 to 3,900,000), the 
hemoglobin from 30 to 50% and the leucocyte count about normal. Such 
cases improve rapidly under treatment. 

Gastro-intestinal disturbances are important causes of secondary 
anemia, and possibly also of many so-called primary anemias in patients 
whose intestinal features were overlooked. The intestinal wall is one of 
the most important of the hematopoietic organs since it is the source of 
supplies for the plasma, hence indirectly for the cells. 

In 60% of our cases of severe diarrhea in men the red count was not above 4,000,000; 
in women it was a little higher. The real anemia must have been more pronounced than 
this for in cases of diarrhea the blood is concentrated by the loss of fluid ( in one case 
the count was 7,900,000). The leucocytes often ran low (even to 2700 and 2500) 
but in 30% of the cases the count was above 10,000. 

Cabot mentions 1 such case with 1,928,000 reds, and another with 2,440,000 and 
10% hemoglobin 

In chronic dysentery the red cell-count may be high or low. In 1 case it was 1 ,520,000, 
in another 2,500,000. On the other hand, in 1 (a male) it was 7,000,000 with 1 10% hemo- 
globin and 7000 leucocytes; and in another (a woman) it was 6,300,000. 

In chronic constipation our cases showed both normal and high counts. 

Our cases of benign dilated stomach showed nothing abnormal as regards the leuco- 
cytes. The red counts were low yet within normal limits, except in 4 cases which showed 
considerable anemia (3,300,000, 2,400,000, 2,250,000 and 2,600,000). The vomiting of 
large amounts of fluid does not seem necessarily to concentrate the blood. 

Acute gastritis, during the febrile period, causes a slight leucocytosis and this was 
true also of 70% of our cases of gastro-enteritis. A slight leucocytosis is common also in 
chronic gastritis, except the alcoholic form in which cases the counts were quite low. 

In ulcerative colitis the red cell-counts below 3,000,000 are not rare. 

In amebic dysentery a severe anemia is rare, yet in 24% of our cases there was a slight 
(4,000,000 to 4,500,000) and in 12% a more severe (2,200,000 to 4,000,000)' anemia. 
A leucocytosis was the rule (in 70% of cases) at some time during the disease, the highest 
count being 19,200. Futcher U3 found the general average of 43 cases about 10,000. 
In children Amberg found an eosinophilia. 

Anemia op the Tropics. — It is said that Europeans after a stay of 
some duration in the Tropics look anemic. Some consider this only ap- 
parent although the frequent presence of basophile granulations in the 

113 Jour. Am. Med. Assoc, August 22, 1903. 



THE BLOOD 599 

red blood-cells would seem to indicate some injury to these cells. There 
are several tropical diseases, important causes of anemia, which only 
now we are beginning to understand and these may explain some of the 
above cases. 

Chronic Infectious Diseases. — Of the chronic infectious diseases 
three are most potent causes of anemia, — lues, tuberculosis and leprosy. 
While the toxins of the organisms of these diseases may be important yet 
the gastro-intestinal conditions, the lack of exercise and the frequent hem- 
orrhages so common in these diseases also must be considered. 

There is a great difference in the effect of bacterial toxins on the blood. 
In acute miliary tuberculosis without cyanosis, for example, i of the worst of 
septicemias, there is little trace of blood destruction (see page 637), while 
anemia, is a common result of latent pyogenic infections. The same is true 
in diseases with chronic exudate formation. Albuminuria is frequently cited 
as an important cause of anemia and yet the anemia must be due to the 
cause of the albuminuria for the actual daify loss of proteid to the blood- 
plasma even in a severe case is very slight. 

Spermatorrhea, lactorrhea, and diseases of the respiratory organs with a 
large amount of sputum are given as causes of anemia, and yet patients with 
long-standing profuse purulent exudate formation, as chronic bronchitis 
and tuberculous abscess of its joints, maintain their blood condition 
surprisingly well. 

In cases with marasmus an atrophy of the total blood volume may cover 
an anemia, while in other cases the anemia may be more apparent than 
real, since because of improvement the plasma is diluted. 

Fever is cited as an important cause of anemia and yet it is not the 
elevated temperature but the toxins which cause the rise of temperature 
that destroy the red cells. Most important are those cases of chronic 
cryptic septicemia which for weeks may present the picture of a severe 
anemia. Acute infections may cause a rapid fall in the blood-count, as for 
instance Grawitz's case of streptococcus septicemia, in which in a little 
over 1 day the reds fell from normal to 300,000. 

A recent case of arthritis of unknown cause, but with blood-cultures 
negative, had a red cell-count of 976,000, Hb 17%, and leucocytes, 4600. 
He improved rapidly. 

In yellow fever considerable anemia may develop. In 1 case the count 
was 2,604,000 and in another 1,400,500 (Maurel). 

Pneumonia, diphtheria, scarlet fever, typhoid fever, acute articular rheu- 
matism, smallpox, septicemia and other acute infectious diseases may cause 
a severe anemia. The reader is referred to the various sections on these 
diseases. In all such cases there may, for the first few days at least, be no 
diminution in the red blood -count, but rather a hypercythemia due to con- 
centration of the blood. This is best seen in diphtheria and typhoid fever 
and it may cover a real anemia. The rapid fall in the count in cases of 



600 CLINICAL DIAGNOSIS 

pneumonia during convalescence or at the time of the crisis, is probably 
more apparent than real and due to dilution of the blood resulting from a 
temporary general vasomotor relaxation (Grawitz) . 

In many cases of infection there is merely a drop in the count, but in 
very severe cases microcytes, macrocytes or poikilocytes are present. In 
these latter cases hydremia is the rule, the loss of albumin running parallel 
to the severity of the disease, and in severe cases reaching even 6.25 gms. 
of residue per 100 c.c. of blood. 

Intestinal Parasites. — Of the intestinal parasites 2 are very impor- 
tant causes of anemia. 

Uncinaria Duodenalis et Americana. — Historically this form of anemia 
is most interesting since the miners' and tunnel diggers' anemia due to this 
parasite were the first forms of anemia accurately to be described. This was 
then rated as primary pernicious anemia. 

These parasites are met with in many different countries and in this 
they would seem to be the chief cause of the "anemia of the South." One 
severe case we saw had a red cell count of 2,424,000, Hb 32% and leucocytes 
9700, of which the eosinophiles were 5.6%. Counts below 1,000,000 cells 
have been reported. 

This anemia resembles one due to hemorrhage rather than to a toxin. 
(Note the small amount of iron in the liver, even but % the normal 
amount, the absence of a leucocytosis, and the very low color-index.) 

In 3 cases of Strongyloides intestinalis infection the blood showed : in 
the first, red cells 5,420,000, Hb 82% and leucocytes, 6200; in the second, 
3,560,000, 57% and 21,500; and in the third, Hb 60% and leucocytes, 7500. 

Bothriocephalus Lotus. — Bothriocephalus latus (see page 417) may cause 
a most interesting and severe anemia, the almost exact picture, both quanti- 
tatively and qualitatively, of the primary pernicious type, but which will 
disappear soon after the worm has been expelled. In Lichtheim's case the 
count of red blood- corpuscles dropped to 500,000 and the hemoglobin to 
20%. Six worms were expelled. In Schapiro's case the count was 837,000 
and in 23 days after the worm was expelled, 2,975,000. Bezancon and 
Labbe give 1,300,000 as the average of reds in a series of cases, (the limits 
were from 395,000 to 2,150,000) while the color-index varied from 0.9 to 
1.62. All the signs of a severe primary anemia, the poikilocytes, microcytes, 
macrocytes, the polychromatophilic degeneration, etc., are present. Even 
T /2 of the nucleated reds may be megaloblasts and yet in 2 weeks after the 
worm has been expelled the megaloblasts may all disappear, in 3 weeks all 
the megalocytes and soon the blood appears quite normal. The leucocytes 
are normal both quantitatively and qualitatively. 

The cause for this anemia is unknown. It is not the mere presence of the worm, 
since but 16% of the persons who harbor this parasite have any anemia. It is not the 
duration of the infection, for some persons are hosts for even 20 years before the anemia 
begins. There is no evidence of hemorrhage and the amount of iron of the liver has been 



THE BLOOD 601 

found even twice normal, which would indicate an intravascular destruction of the red 
blood-cells. Schaumann believes that the patients must have some predisposition to 
an anemia. Dehio says that the worm itself must become diseased or die to affect the 
host's blood. But in some cases with anemia the worm does not appear diseased and in 
others who harbor a certainly diseased worm there is no anemia, while in still others 
the anemia persists after the worm has been expelled. 

Tcenia saginata and Tcenia solium, Strongyloides intestinalis (in cases of 
the "diarrhea of Cochin China" due to this parasite, counts as low as 
760,000 have been reported) and Ascaris lumbricoides, sometimes, it is 
claimed, cause anemia. 

Yeasts. — In four cases of systemic blastomycosis a moderate secondary 
anemia has been reported (e. g., 3,992,000 red cells per c.mm.), and in 8 cases 
the leucocytes varied from 9,600 to 2 1,200. In 2 of these cases this yeast was 
isolated in pure culture from the blood. 114 In 1 case of infection with 
Oidium coccidioides there was a leucocytosis of 17,000. 115 

Poisons. — Lead, mercury, arsenic, certain organic poisons, plant and 
animal toxins, ptomaines and the toxins of burns, all may produce an 
anemia. Lead is a very important cause of both the acute and chronic 
forms of anemia. It causes essentially a chlorotic anemia, manifested 
first by degeneration (?) of the red blood-cells, not by any diminution in 
number, after which the count is reduced even to 1,300,000. Megaloblasts 
sometimes are found. The basophile granules of the red cells are early 
very common and important in diagnosis (see page 479). 

In the Johns Hopkins clinic there were 17 cases of anemia due to lead. Of 16, the 
lowest red count was 2,900,000; in 7 the red count was over 4,500,000; the mean was 
4,200,000. The lowest hemoglobin was 38%; the mean, 60%. In 10 of 16 cases the leu- 
cocytes were above 10,000, with the maximum 25,000. This count fell very soon after 
admission. 

Long-continued use of certain of the coal-tar products will cause a severe 
anemia. Stengel and White 116 report a most interesting case due to ace- 
tanilid in a woman whose red cell count was 2,092,000, Hb 35%, and 
leucocytes 19,800 (a previous count). There were 32,323 nucleated reds 
per cubic millimeter, of which 91.4% were normoblasts, 3.5% megaloblasts 
and 5.3% free nuclei. The number of platelets was increased. Many of the 
red cells were poikilocytes, a few contained basophile granules and many 
showed polychromatophilic degeneration. The diagnosis in this case was 
suggested by the appearance of the smear alone. They mention Ehrlich 
and Lindenthal's case in which 1 of each of 56 red cells was nucleated. In 
Brown's case of acetanilid poisoning 117 the reds at death numbered 
1,166,000, of which 22,150 per c.mm. were nucleated. 

m Montgomery and Ormsby, Arch, of Int. Med., Aug., 1908, vol. ii, No. 1, p. 1. 

115 Hektoen, Jour, of A. M. A., Sept. 28, 1907, vol. xlix, p. 1071. 

116 Contrib. of the Wm. Pepper Laboratory of Clinical Medicine, 1903, No. 4. 

117 Amer. Jour. Med. Sci., 1901, vol. cxxi. 



602 CLINICAL DIAGNOSIS 

Splenic anemia is the name given to a group of cases of anemia of the 
secondary type associated with idiopathic enlargement of the spleen. The 
count in Osier's cases averaged over 3,000,000. The leucocyte count was 
normal or reduced. Such cases suffer profuse hemorrhage from gastric and 
esophageal varices. In 1 of Osier's cases the macrocytes and gigantoblasts 
were a marked feature. 118 

Simple Primary Anemia. — Some authors attempt to separate a simple 
primary anemia from the primary pernicious form, basing- their differences 
on the clinical course which is characterized by the number of relapses 
and which ends finally in death. It would be hard to separate this dis- 
ease from the secondary anemias already mentioned as due to bad 
unhygienic conditions. These cases seem to stand midway between chlor- 
osis and primary pernicious anemia, with oligocythemia and oligochrom- 
emia of about equal grade and with leucocytes normal both quantitatively 
and qualitatively. 

Progressive Pernicious Anemia. — Eichorst's definition of primary per- 
nicious anemia is, a severe anemia which in spite of all treatment pro- 
gresses relentlessly to death. Many cases, however, live for years with 
occasional remissions and intermissions. It is usually referred to as a 
hemolytic anemia. Clinically the blood picture is very definite although 
not characteristic. Pathologically there is evidence of an excess of broken 
down hemoglobin and bone marrow lesions which are almost character- 
istic; that is, a hyperplasia of red marrow containing an unusual number of 
megaloblasts. The blood picture alone is not characteristic, for many cases 
of definitely secondary anemia have a similar picture, e. g., the "secondary 
pernicious anemia" which develops in rare cases of cancer, phthisis, lues, 
malaria, following repeated hemorrhage, lead poisoning, certain animal 
parasites, lesions of the bone-marrow, especially tumors, also osteomyelitis, 
atrophy of the gastric mucosa, stenosis of the pylorus, nephritis, certain 
rare cases of pregnancy and purpura hemorrhagica. Many of these cases 
give a history of anemia-producing conditions which for years may have 
led to almost complete bankruptcy of the blood-building function. The 
only difference between these cases and primary pernicious anemia is the 
absence in the latter at autopsy of any of the lesions of these diseases. 

The salient characteristics of the blood in primary pernicious anemia are: 
A megalocytic blood picture including megaloblasts ; anisocytosis ; poikilo- 
cytosis ; a high color-index (because of the megalocytes) ; marked evidences 
of blood regeneration (nucleated red cells of all types) ; and signs which 
have been interpreted as those of rapid blood destruction (degenerated 
(?) reds, endoglobular degenerations, polychromatophilia, urobilinuria, 
jaundice, the increased iron compounds (?) in the serum and the corpuseles, 
and the increase of iron stored in the liver and spleen) . The student should 

118 Osier, Am. Jour. Med. Sci., January, 1900. 



THE BLOOD 603 

remember that the presence of all this pigment need not indicate in- 
creased hemolysis but the effort of the body to get rid of hemoglobin which 
it can no longer use. There is a constant and normal breaking down of 
cells which have served their purpose (at least 18 gms. of hemoglobin per 
day and probably more) and if the bone-marrow is unable because of def- 
inite disease to build new cells fast enough some of this pigment must be 
destroyed. At first the poikilocytosis was supposed to be characteristic 
of this disease (Quincke) but this idea was very soon corrected; then the 
high color-index (Laache and Kahler) and this is still the opinion of many ; 
then the presence of megaloblasts (Erhlich) but this is not true, although 
they are particularly numerous in these conditions. The diagnosis cannot 
be made from any one feature. 

Volume of the Blood. — There is no satisfactory clinical method of de- 
termining blood-volume but in pernicious anemia the appearance of the 
patient suggests and later the autopsy shows a great reduction in the 
amount of blood in the heart and blood-vessels. In a remarkable case seen in 
Professor Miiller's clinic all the organs seemed almost exsanguine. 

Gross Appearances. — The ear is a better place to obtain a drop of blood 
than is the finger. It may flow freely or not at all. Lazarus considers that 
the former occurs when the patient is doing badly and that the latter is 
evidence of improvement. The blood is pale, of a light red, watery color 
and may not at all resemble blood. 

We held up a tube full of this blood before a class on one occasion and asked them to 
tell from its appearance alone what fluid it was. Many of them said it was a cloudy 
urine, which, indeed, it did resemble. 

The drop of blood is often grossly streaked, evidence that the corpuscles 
have collected in masses. Cases have been described in which it grossly 
is of a coffee-color, probably due to hemoglobinemia. The coagulation 
time is often increased. 

Red Blood-cells. — In the fresh specimen the poverty of the blood in red 
corpuscles and the absence of rouleaux formation are conspicuous features. 
The cells vary much in size (anisocytosis) ; the majority are slightly above 
normal, some are very large and others are small, some very small. A few 
show Maragliano's endoglobular degeneration and a few more show another 
degeneration, the accumulation of the hemoglobin in the center of the cell, 
but the most have a uniform dark color. Nucleated reds often are nu- 
merous. In a well-marked case the appearance of the fresh specimen alone 
will strongly suggest the correct diagnosis. 

An extreme oligocythemia is the rule in pernicious anemia. It is re- 
markable how few symptoms the patients with these low counts suffer, 
particularly as the volume of blood also is dimished. On the first visit the 
average cases will show a count of about 1,000,000 cells. Cabot's average 
was 1,200,000. 



604 CLINICAL DIAGNOSIS 

In the 8 1 cases of the Johns Hopkins series which we studied the average red cell- 
count on admission was 1,575,000. This is somewhat higher than that which other 
observers have reported, since we included in the series patients admitted for complica- 
tions of the disease and not for the asthenia of the anemia alone. In 81% of these cases 
the count on admission was under 2,000,000 and in 12% under 1,000,000. 

Some patients with counts as low as 500,000 are comfortable and active, 
while others with 4 times that number of cells suffer severely. Evidently 
the reason for their symptoms is not the oligocythemia alone and yet the 
first counts which the same patient has on several successive admissions are 
often curiously similar. The count may remain stationary for some time, 
often in the neighborhood of 1,000,000, or it may diminish progressively 
until death. Quincke reported 1 case with a blood-count of 143,000 who 
recovered. Hayem reported a fatal case whose lowest count was 292,000. 
Scott's case at death had 268,000 red cells, a color-index of 2 and 5900 
leucocytes per c.mm. ll9 

After treatment begins the count may continue to drop for a while and 
then rises ; or it begins to rise at once. In a few cases it remains stationary, 
but as a rule there is a tidal rise and fall of the count which is important 
in diagnosis. 

It cannot too often be emphasized that a change in count may mean a 
change in the total number of red cells or a change in the volume of plasma 
and that improvement is usually ushered in by a drop in the count due to an 
increase in the volume of plasma. 

It is interesting how few clinical symptoms seem related to the red blood cell-count. 
Judged by the comfort of the patient alone a case of pernicious anemia with a count of 
1,000,000 is more comfortable than a case of chlorosis with a count of 4,000,000. Again, 
in some of those cases in which the count continues to fall after admission and then later 
rises, it is curious that the patient feels so well that he insists upon going home at a time 
when the count is no higher or very little higher than on his admission. In other cases 
in which the count rises after admission and then falls, death occurs when the count 
has reached about the level of admission. In still other cases with an initial drop, as in 
5 of our series, the count was rising at the time of death. 

The red blood-count on the day of death of 2 of our cases was high — 2,700,000; in 
3 cases it was moderate, 1,031,000, 1,326,000 and 1,216,000; while in 13 cases, and this, 
we think, is a hint of the blood picture if death is due to this anemia alone, the count 
was between 718,400 and 376,000, an average of 567,700. 

The blood during the intermissions of this disease is not quite as normal 
as the subjective feelings of the patients would suggest. The red count 
averages about 3,000,000, the cells are still a little large (Cabot), the nu- 
cleated cells have disappeared, the color-index is usually high (but some- 
times low) and the leucocyte count is increased by an increase in the poly- 
morphonuclears. The diagnosis of this disease during intermissions is 
important, especially to insurance examiners. Two of our cases with almost 

119 Am. Jour. Med. Sci., 1903, vol. cxxv., p. 397. 



THE BLOOD 605 

normal blood (i was refused by i examiner) both succeeded finally in getting 
heavy insurance and died in about i year. 

The volume of the red blood -cells, which is determined better by sedi- 
mentation than by centrifugalization, averages from 8 to 10%, which is 
high relative to the count and is a measure of the large size of the cells. 
Capps found that in this disease the volume index is always higher than the 
color-index. The cells are little biconcave (seldom does one see pessary 
cells) but seem plump although shadows do occur. 

The average diameter of the red cells in primary pernicious anemia is 
definitely increased, although the variations in size are very wide, in general 
from 4 to 13^, but with extremes beyond these. In no other disease are 
there so many macrocytes and also microcytes but the mean size is definitely 
increased (a few cases are mentioned in which the blood is not megalocytic) . 

Seventy per cent, of the cells are macrocytes and measure between 1 1 
and 13/x in diameter (Lazarus) . In a case reported by Ewing 90% measured 
from n to 16 p. Gigantocytes are numerous. These huge cells are not 
biconcave, sometimes are biconvex, often are oval and seem flabby. Many 
appear dark in fresh and polychromatophilic in stained specimens. A few 
may be pale (Grawitz) ("chlorotic cells," "dropsical cells"), but the dark 
color of most of them is a quite constant feature. Some cells seem to have 
a slight change of color shade as well as of color depth. The presence of ma- 
crocytes is considered (Laache) a compensatory attempt to replace the 
amount of hemoglobin-containing protoplasm; Cohnheim first said it was 
reversion to the embryonic type; Ehrlich attributes it to a megaloblastic 
degeneration of the bone-marrow (see page 502). 

Microcytes, cells which vary from 2 to 6/jl in diameter, occur in large 
numbers here as well as in the secondary anemias. So numerous may they 
be that the average size of the red blood-cells may not be above normal. 
Their dark color may be due to their spherical shape, but they have some- 
times a greenish tint, suggesting a chemical change in the protoplasm. 
These microcytes change their shape definitely; they also move quite ac- 
tively among the other cells with an oscillatory motion which suggests 
ameboid activity. They have been described as monads, a leptothrix 
form, bacteria, and Hayem called them pseudo-parasites (see Fig 117). 

Poikilocytes, or deformed cells, are present in great numbers and in a 
great variety of shapes : hooks, raquette forms, spindle and various dwarf 
forms (microcytes, see above), but it is the sausage and battledore shapes 
which, it is claimed, are found here only. Since in an acute case abnormal 
cells did not appear until about 2 months from onset, McCrae suggests that 
such cells are products not of degeneration but of defective regeneration. 

Polychromatic "degeneration" is best studied in pernicious anemia. 
Despite the name many of these cells may be young forms. With Ehrlich 's 
triple stain they are a pale gray (Plate I, 25-28), while with methylene blue 
they take a blue tint. Their number seems almost parallel to the severity 



606 CLINICAL DIAGNOSIS 

of the case (Grawitz). Red cells with Grawitz's basophile granules are very 
common, especially in severe cases, and have, Grawitz thinks, an important 
diagnostic value. 

Nucleated Reds. — Normoblasts (Plate I, 29, 30, 35, and Fig. 124, a, c, 
d, e), described on page 500, occur quite constantly in pernicious anemia, 
alone or with megaloblasts, and in especially large numbers during the blood 
crises. In a case of Bezancon and Labbe there were^from 6000 to 10,000 
normoblasts and 960 megaloblasts per cubic millimeter. _ Ma.ny of these 
cells show polychromatophilic degeneration, especially those in which the 
nucleus is dividing. 

The blood crisis, so interesting a feature in cases of severe anemia (see 
pages 501 and 608), is not always, as v. Noorden thought, (except in young 
patients), the sign of a regeneration active enough to be followed by a 
rise in the red count, although in secondary anemia and chlorosis this 
usually is the case. The crises probably indicate an attempt of the bone- 
marrow to replenish the blood, but these attempts often are futile and sug- 
gest a convulsive attempt to stem the tide of the anemia. 

In some cases there are few or no nucleated reds in the peripheral blood. 
This means a slower regeneration. In other cases just before death all these 
nucleated cells disappear. 

Megaloblasts (Plate I, 32, 33, 38, and Fig. 124,/) were first described by 
Ehrlich as characteristic of pernicious anemia. They may, however, occur 
in any form of anemia and in any disease which involves the bone-marrow. 
Since there has been little agreement as to the definition of a megaloblast 
the literature on this subject is much confused. To some a megaloblast 
is any large nucleated red; to others it is a red cell with a large nucleus; 
while others describe it as a red cell with a characteristic nucleus irrespective 
of the size of the cell. In the blood of these cases one finds large red cells 
with nuclei like those of normoblasts and smaller cells with nuclei like those 
of megaloblasts. We count both these as intermediate cells, and reserve the 
term megaloblast for a large red cell whose nucleus is at least the size of a 
normal red blood-cell; that is, 7m in diameter. These cells are round or oval 
and vary from about 1 1 to 20 /jl in diameter. The very large ones are called 
gigantoblasts. They are plump, often diffluent, stippled and polychromato- 
philic. The nucleus is plump, round or oval and often surrounded by a 
clear circle outside of which the protoplasm often stains deepest; karyo- 
kinetic figures are sometimes seen which to some is a grave sign. In the 
fresh specimen these nuclei have a well-defined chromatic network, but 
often stains so faintly that it may be overlooked. The fresh unstained 
specimens should be studied whenever possible since it is no easy matter 
to tell in the stained specimen a polychromatophilic megaloblast with a 
palely staining nucleus from a mononuclear leucocyte; indeed it may be 
impossible (Plate I, 36). Color, in a stained specimen, counts but little for 
there is no staining reaction which is characteristic of hemoglobin especially 









THE BLOOD 607 

when basophilic. It is of assistance, however, that the protoplasm of 
the red cell is more opaque, that its margin is thick, smooth, uniform 
and rounded (best seen when this cell touches another), while the 
spherical leucocyte which has flattened out in the preparation has a 
thin, frayed margin. 

While megaloblasts may be found in any anemia they are quite common 
in this and yet they may be hard to find. Ehrlich, it is said, would hunt for 
hours until he found this much-desired cell upon which he would base his 
diagnosis. In some cases for a while none will be found and later, many. 
Their presence in large numbers is an ominous sign. They are some- 
times present during the periods of intermission while the blood -count is 
almost normal. 

Typical megaloblasts are found most frequently in pernicious anemia, 
even mild cases, and for the diagnosis of this condition in adults they are of 
great value. In the various anemias of children, however, one frequently 
meets with them. In adults they may be numerous in splenomyelogenous 
leukemia, in bothriocephalus anemia, in malignant disease of the bone- 
marrow and in malaria, especially of children, even when there is no marked 
anemia; that is, in those conditions in which the bone-marrow is involved. 
Their presence may be an indication of the severity of an anemia, of its 
form, or of bone-marrow involvement. Seme have described them as 
swollen, hydremic cells which contain the increased water contents of the 
plasma. If so they would be similar to the so-called "chlorotic cells" 
seen in cases of marked hydremia, as nephritis, but also in chlorosis, in 
which disease the plasma is practically normal. But these they do not 
at all resemble. 

Intermediate Forms (Plate I, 31, 37, and Fig. 124, b) are those nucleated 
red cells too large, and with a nucleus too large, to be called normoblasts 
and yet not typically megaloblasts. 

That such cells could be transitional between megaloblasts and normo- 
blasts was denied by Ehrlich and Pappenheim but claimed by others 
(Schaumann). They are seen only in those conditions in which megaloblasts 
would be expected, and have, we believe, practically the same significance 
(see page 501). 

Microblasts are cells with a nucleus like that of normoblasts, although 
usually more pyenotic, but with protoplasm exceedingly scanty and often 
ragged on the margin. Whether these cells are normoblasts which have 
undergone some degeneration or are a definite type of cell is not yet known. 
They are met with in pernicious anemia but also in severe secondary, espe- 
cially the post-hemorrhagic, anemias. 

The presence of nucleated reds was noted in 57 of 69 of the Johns Hopkins series 
of cases of pernicious anemia. In 13 definite blood crises were present; that is, there 
were more than 50 nucleated reds per 1000 leucocytes. This is rather an arbitrary line, 



608 CLINICAL DIAGNOSIS 

and yet we have found that, in these cases at least, it corresponded quite well with the 
blood pictures. In all cases some of these cells were normoblasts while in 40 (58%) 
megaloblasts were also present. In the other 6 cases the nucleated reds were normoblasts 
and intermediate forms. 

During the course of these 57 cases were 63 periods with nucleated reds present. 
Of these 26 (that is, 41%) were followed by a gain in the red cell-count; the rest by either 
no gain or by a loss. Of 14 periods with nucleated reds absent, 8 were followed by a 
distinct gain. 

In 13 of these cases (19%) blood crises developed. Five of these cases died. Of the 
50 or more nucleated cells per 1000 leucocytes which constituted the crises, the normo- 
blasts varied in number from 5 to 3128; the intermediates reached even 212 and the 
megaloblasts 44. There seem to be 2 definite forms of blood crises ; those in which normo- 
blasts largely predominate and those in which the intermediate and megaloblasts also 
are present in considerable numbers. The normoblastic crises particularly are followed 
by a rise in the red count; those with many megaloblasts present would seem to be less 
efficient and to appear especially when the patient is losing ground. 

The most remarkable blood crisis of our series lasted for 19 weeks, during which 
time the red blood-cells, at the beginning 1,902,000, rose to 2,562,000, and then at death 
had dropped to 1,328,000. The leucocytes, meanwhile, varied from 3000 to 5000 until 
the day of death when the count was 16,000. During this whole period the blood con- 
tained over 500 normoblasts per thousand of leucocytes, on one occasion 1164, a little 
later 1032, and once 3128. Since on this last day the leucocyte count was 4600 the total 
number of normoblasts must have been 14,388, the intermediate forms 460 and the mega- 
loblasts 138 per cubic millimeter. 

Hemoglobin. — The hemoglobin is much reduced in pernicious anemia. 
It is rarely above 50% and often is as low as 10%. The color-index, how- 
ever, is normal or high, a point of great importance in diagnosis. In our 
cases the hemoglobin on admission averaged 34% and the color -index 1.1. 
In 80% of the cases, it was over 1 and in 2 cases as high as 1.9.(0 

Ewing considers that a low index indicates a chronic case and a high 
index an acute one. For it to rise is considered a bad sign since it indicates 
a falling count ; with improvement there is always a lowering of the index 
since so many of the newly formed cells are of lighter weight than normal. 

The high color-index in pernicious anemia has received various explanations. One 
is that it is the result of abnormal " globular richness "; that is, of an abnormally large 
amount of hemoglobin per cell. This idea is confirmed by estimations made of the weight 
of the cells and by Capps who found that the color-index never exceeds the volume-index. 
Another explanation is that it is due to the large number of macrocytes present and 
indeed the hemoglobin curve does run fairly parallel to that of the number of these large 
cells. Others ascribe it, and with good reason, to incorrect blood counts in which a 
great many of the microcytes have been overlooked, while their hemoglobin does add 
to the color test. Others say that in pernicious anemia some of the hemoglobin is free 
in the plasma. Some believe that the chemical composition of the red blood-cells in 
this disease is not normal and find their nitrogen increased (v. Jaksch) , hence the name 
" hyperalbuminemia rubra." Others find more iron present than theoretically the hemo- 
globin molecule should contain, which would mean either that iron is increased in the 
hemoglobin molecule, or is present in other combination. In favor of this is the hema- 
togenous jaundice so often present and the discovery reported of iron compounds in the 
plasma. Taylor considers the high color-index an optical illusion; Grawitz (and this 






THE BLOOD 609 

view appeals to us very strongly) warns against hemoglobin determinations with an 
ordinary hemoglobinometer and emphasizes the danger of overlooking microcytes in 
blood-counting. He considers that the index is best determined by the appearance of 
the cells and thinks that the inequality in the distribution of protoplasm and the produc- 
tion of poor cells are the prominent features of pernicious anemia. Bezangon and Labbe 
say that the appearance of the cells does not suggest that they are overrich in hemo- 
globin. It is our opinion that it does (see page 501). That changes in the composition 
of hemoglobin occur and can make red blood-cells appear darker than normal is illus- 
trated by many degenerating cells, by cells picked up by phagocytes and by the brassy 
cells of malarial blood. 

It is important to remember that the hemoglobinometers with a color prism do not, 
as bought, give accurate readings in the lower half of the scale unless they have been 
especially standardized and that an error of 5%, so insignificant in normal blood, changes 
the index considerably when added to a total of but 10%. 

During the course of our cases the hemoglobin, as a rule, ran parallel with the count 
of red blood-cells. As the cases became worse the index slowly rose and at death averaged 
1.5. This may have been due to the tendency of the marrow to form large cells. 

Leucocytes. — In severe and uncomplicated cases of pernicious anemia 
there is always a leucopenia. Cabot's average count was 3800 and in 72 
of the no cases it was below 5000. The leucocytes in our cases on ad- 
mission averaged 4600. This includes all cases, even those with leucocytoses 
due to complications. In 75% of these cases the count was under 5000. In 
some it went as low as 2000 or 1500 and before death even to 500 cells per 
cubic millimeter. The leucocyte count runs, as a rule, parallel to that of the 
red blood-cells. A leucocytosis may mean a complication, as pneumonia, 
a pyogenic infection or a blood crisis, in which case the large number of 
leucocytes may even suggest leukemia. At death, the picture may be 
leukemic (100,000). 

The leucocyte count in our cases ran fairly parallel to the red count. At death it 
varied from 660 to 16,000 and averaged 5950. 

Of our 81 cases, in 55 (70%) this count fell at some time during their stay below 3000; 
in 32 (40%) below 2000; in 9 (n %) it fell to 1000 or below. These very low counts, 1000 
or below, are found only in the severest cases. 

The percentage of polymorphonuclear neutrophile cells runs roughly 
parallel to the total leucocyte count. This is well illustrated by the rise of 
these cells with the improvement of the case and their low percentage when 
the count is low. 

The absolute number of the mononuclear non-granular cells tends to be 
quite constant, therefore their percentage will vary inversely with that of 
the granular cells. The highest per cent, in our cases was 93%. It is also 
true that in any given case the percentage relation of the various types of 
leucocytes tends to be constant whatever the total count of these cells, 
which indicates that some of these variations are due more to the distribution 
of cells or to dilution of the blood, perhaps from stasis in the vessels, than 
to any real change in the blood formula. 

Toward death the percentage of the mononuclear non-granular cells 
39 



610 CLINICAL DIAGNOSIS 

rises, probably because the granular cells are then being formed in diminish- 
ing numbers. It is interesting that the variations in the percentage of these 
cells show definite waves during the course of the disease . 

The percentage of lymphocytes is always high, averaging 45%. This 
seldom indicates a true lymphocytosis but rather an absolute decrease in the 
number of polymorphonuclear cells. Their percentage may reach as high 
as 62 and before death even 79%, and yet their absolute number remain 
normal. This has been given as evidence that these cells arise in the lymph- 
glands and not in the bone-marrow. This decrease in the number of poly- 
morphonuclear cells is a striking feature in pernicious anemia and in diag- 
nosis is used to exclude cancer and septic anemia. Yet even in the same 
case these cells vary so much that this point is not of great importance. 

Twelve (17%) counts indicated a true lymphocytosis which was temporary in every 
case. In one there was a definite leucocytosis, while in the others the total counts were 
not above normal limits. 

The eosinophiles averaged about 2.7%. In 1 case they reached 9%. They are 
often absolutely increased, but in severe cases are more often diminished. 

Myelocytes are often found. In no disease except leukemia are they 
so constantly present or their number so great as in pernicious anemia. 
In acute exacerbations of this disease they may make up even 29.4% of a 
total of 34,000 cells (Billings). Eosinophilic myelocytes sometimes, but 
rarely, are met with. 

In 23 of our cases myelocytes were present in numbers varying from 0.2 to 8%. 
Nine of these cases were fatal. In 12 cases the percentage was not above 1%. In 6 it 
was above 3%. 

In our cases the myelocytes were conspicuous under 2 conditions; in cases with a 
very low count and when a leucocytosis was present. In 1 case the total leucocyte count 
was 1800, 8% of which were myelocytes. In another case with 14,400 leucocytes 2% 
were myelocytes. In another case the leucocyte count was n,6oo of which 0.5% were 
myelocytes. 

Mastzellen were noted in 29 of our 69 cases. In 2 the}?" made up over 3% of the 
total count and in 8 over 1%. The average total leucocyte count in these 8 cases was 
3900. Of the other 21 cases the average per cent, of Mastzellen was 0.5%. If in these 
cases the cells thus designated really were Mastzellen, then the blood in pernicious anemia 
shows a definite increase of these cells hitherto not mentioned. 

Many of the leucocytes in pernicious anemia show evidence of degen- 
eration; that is, are pale, swollen and vacuolated and their nuclei fibrillar. 
The neutrophile granules of some cells are crowded at the periphery. 
Hayem considers that some imbibe a certain amount of hemoglobin. 

Some cases of pernicious anemia resemble closely acute leukemia. In 1, 
for instance (Williamson and Martin), the red blood-cell count was 300,000, 
the hemoglobin 12% and the leucocyte count 38,000 of which 99% were 
small mononuclears. In Westphal's case the red cell count was 816,000 
and the leucocyte, 24,000. In Bezaneon and Labbe's case there were 



THE BLOOD 611 

500,000 red cells per cubic millimeter of which 3250 were nucleated, and 
32,000 leucocytes of which 66% were small mononuclears. 

The curve of the absolute number of eosinophiles runs - in many cases 
parallel to that of the red blood-cells. During sixteen admissions a definite 
rise of these cells accompanied some improvement of the condition. The 
coarsely granular cells, therefore, have considerable value in prognosis since 
their count is not affected by changes in the volume of the plasma. The 
highest counts of these cells were found chiefly in those cases which are 
certainly doing well or after they have already done well. In several cases 
in which the general condition of the blood changed little the number of 
eosinophile cells remained fairly constant. In 10 cases these cells fell as the 
red blood-cells dropped and in 3 they had entirely disappeared at the time 
of death and had almost in 2 others. These cells may diminish as the pa- 
tient's condition gets worse even though the red count does not. On the 
other hand in 8 there was no rise in the count of eosinophiles as the red 
count rose (plasma changes ?) while in 4 there was a rise but without any 
accompanying improvement. In 1 case, a terminal pneumonia, these cells 
numbered 220 at death. It may be noted that in the cases with apparent 
improvement but with no rise of eosinophiles the increase in the count of 
red cells averaged 43,000 per day; while in cases with definite improvement 
in the blood condition and with a definite eosinophilia, the average gain 
of red cells per day was 17,000, i. e., was slow and lasted over a considerable 
period of time. This would suggest that in the latter cases there is real new 
formation of blood while in the former the rapid changes in the red count 
may mean merely plasma changes. The largest counts of eosinophiles were 
seen in those cases which were gaining very slowly but surely. 

A diminution in the absolute number of eosinophile cells may be of ill 
omen. In 1 of our fatal cases, for example, the red cell-counts had for 15 
days before death remained constant (that is, the first of 6 counts was 
2,832,000, the last 2,704,000, and the average in all was 2,700,000). The ab- 
solute number of eosinophiles, however, at first was 183, shortly afterwards 
was 180, while toward the end none of these cells were found. 

Platelets. — In pernicious anemia the blood-platelets are decreased often 
to only % their normal number, while they may be apparently absent. 
In other cases they are said to be increased (v. Limbeck and Sahli) . Grawitz 
found that they varied. Hay em found the count as low as 25,000 or even 
15,000 per c.mm. 

The coagulability of the blood in pernicious anemia is usually decreased. 
The blood obtained by venesection does not separate into clot and serum. 

Serum. — The plasma in pernicious anemia loses very little of its albumin. 
There may be a loss of 50% of the albumin of the whole blood and yet the 
serum will lose but 8%. Pernicious anemia is quite different, therefore, from 
the hydremic anemias after hemorrhage and those due to cancer, sepsis 
(cryptogenetic infections), etc. 



612 CLINICAL DIAGNOSIS 

The specific gravity of the blood averages about 1.030 and may go as 
low as 1.025. 

The solids of the blood average but about 9% and the water is increased 
to even 90%. The loss is in the albuminous bodies and due to the reduction 
in the count of corpuscles, for the serum in even the severe cases may be 
practically normal. In the plasma the serum globulin alone is decreased 
while the serum albumin remains practically normal. 

The blood lipoid values in anemia were found by Bloor and MacPher- 
son 120 to be "normal, or nearly so, as long as the percentage of blood cor- 
puscles remained above half the normal value. When the percentage was 
below this level abnormalities appeared which, in the order of their mag- 
nitude and also of the frequency of their occurrence were, (1) high fat in the 
plasma, (2) low cholesterol in the plasma and occasionally in the corouscles 
and (3) low lecithin in the plasma. 

"The lipoid composition of the corpuscles was found to be normal in 
all cases. There was therefore nothing in their composition to indicate 
abnormal susceptibility to hemolysis. 

"Removal of the spleen resulted in increased total fatty acids and leci- 
thin in the corpuscles and of cholesterol in the plasma. The results were 
essentially the same whether the patients had anemia or not. 

" The relation between free and bound cholesterol was found to be within 
normal limits in all cases of pernicious anemia, thus giving little support to 
the assumption that an abnormally great combination of cholesterol as 
ester is a factor in the production of anemia. 

"The low values of lecithin and the high values for fat which were 
generally most marked in those cases where the blood corpuscle percentages 
were lowest are regarded as due to deficient fat assimilation in the blood 
resulting from the lack of sufficient corpuscles to bring about the change of 
fat to lecithin which has been found to be one function of the corpuscles. 

"While the results offer no certain evidence that abnormalities in 
the blood lipoids are responsible for anemia, the low values for choles- 
terol, which is an antihemolytic substance, and the high fat fraction, 
which may indicate the presence of abnormal amounts of hemolytic lip- 
oids in the blood, are possible causative factors of which further investi- 
gation is desirable." 

Chlorosis. — Chlorosis is a disease especially of young girls at puberty 
(hence the desire to relate this disease to some defect of sex hormone for- 
mation) , the essential blood-feature of which is a reduction in the size and 
thickness of the red blood-cells. The amount of hemoglobin is, therefore, 
much reduced, much more than is the count of red cells. These cells show 
practically no signs of degeneration or destruction. There would seem also 
to be a polyplasmia. Chlorosis is the best illustration of an anemia due to 

120 Jour, of Biol. Chem., July, 1917, xxxi, p. 79. 



THE BLOOD 613 

defective hemogenesis. The absence of hemolysis is shown by the poverty of 
the urine in pigment and by the absence of jaundice. 

The blood features in chlorosis which deserve special mention are : the 
remarkably uniform diminution in the size of the red cells; their almost 
uniform paleness (in the secondary anemias, even of a severe grade, 
the red cells vary widely in size and in color) ; the low color-index (lower than 
in secondary anemias) ; the lymphocytosis usually present ; the infrequent 
appearance of nucleated reds and the increased coagulability of the blood. 
* And yet chlorosis is more a clinical than a blood picture, since this 
latter is well simulated by many secondary anemias. On the other hand the 
clinical picture is so sharp that some speak of chlorosis without blood 
changes (Laache). 

The gross appearance of the drop is very pale, thin, and watery. It 
clots rapidly. 

The count of the red blood-cells is not very much reduced and very low 
counts are rare, and yet "in over 60% of the cases it is under 4,000,000 cells 
at the time of the first visit" (Reinert, v. Limbeck). Thayer's average of 
63 cases on the first visit is 4,096,000; Cabot's, 4,112,000; Graber's, 
4,482000; while Grawitz's cases varied from 3,400,000 to 4,300,000. The 
minimum count of Cabot's was 1,932,000; of Thayer's, 1,953,000, and 
of Hay em's 937,360. Graber, who claimed that in simple chlorosis there 
is no diminution in the count and that a diminution would indicate a com- 
plication, as ulcer of the stomach, cites one case with 5,700,000 cells. The 
color of the red blood-cells suggests a marked diminution of the hemo- 
globin. Their biconcavity is very pronounced, the pessary forms are 
common and they stain very poorly (Plate I, 23, 24). 

The majority of the cells show a quite uniform diminution in size, and 
yet there are present just enough large, pale, so-called "chlorotic Cells" 
to bring the average diameter up to almost normal (they vary from 5.2 to 
1 1.5M in diameter, with an average of 7. 5m)- Many consider that these 
large cells are dropsical; — that is, that they are swollen by the water they 
have imbibed from the plasma. Macrocytes are rare, while rhicrocytes are 
more common. Schaumann and Willebrand say that at the height of the 
disease the smaller cells predominate, but during convalescence the large 
cells. Grawitz, on the other hand, says the cells are largest when the case 
is at its worst; that these large, chlorotic cells, at the height of the disease, 
may be very numerous, may even make up one-third of all the red cells. 

Poikilocytes and degenerated cells are rare except in the severe cases. 
The polychromatophilia present is considered by many to mean youth and, 
therefore, to be a sign of active regeneration (Grawitz). "The granular 
degeneration does not belong to the picture of chlorosis, but means some 
complication." Stengel and Pepper, however, think it is common. 

Nucleated reds are very rare in chlorosis except in the very severe 
cases and during the blood crises. They certainly are much rarer than 



614 CLINICAL DIAGNOSIS 

in the secondary anemias. Those present are usually normoblasts, 
seldom megaloblasts. 

It is the marked reduction of the hemoglobin which is the characteristic 
feature of chlorosis. This may be reduced to even 20%. Cabot's average 
on first visit was 41.2% and Thayer's, 42.3%. The color-index is, therefore, 
low, averaging 0.5, but in some cases it has been reported as low as 0.3. 
Secondary anemias never reach this level. The explanation in chlorosis is 
the small amount of hemoglobin in each cell and the large numbers of small 
cells. The volume of the red blood-cells is just about half the normal. 

The average leucocyte count in Thayer's cases was 8467 and in Cabot's, 
7485. That is, the count is normal although a leucopenia is not uncommon. 
This is important in diagnosis since in the secondary anemias a slight 
leucocytosis is the rule. During convalescence the leucocytes may in- 
crease more rapidly than do the red cells and there develops, therefore, 
even a leucocytosis. 

Grawitz and v. Limbeck say that the blood formula is normal and that 
is the present opinion. Those observers who find, even in mild cases, the 
percentage of small mononuclears about 33% and that of the neutrophile 
cells correspondingly reduced probably still cling to Ehrlich's first formula, 
which now is considered incorrect. Some cells resemble myelocytes, but 
typical ones are very rare. The eosinophile cells are usually somewhat 
increased, averaging 3.5% and in some cases reach even 9.6%. 

Chlorosis is a condition met with much less frequently than a few years 
ago. During 5 years but 13 cases were admitted to the Johns Hopkins 
female wards diagnosed as chlorosis. Of these but 2 were at puberty, and 
the rest were from 17 to 25 years old (relapses?). Of these, the lowest 
count was 2,600,000, the highest 4,000,000 and. the mean, 3,700,000. 
The hemoglobin varied from 26 to 49%, the color-index from 0.36 to 0.63 
(the mean 0.47). The leucocyte counts were 2400 and 3800; between 
5000 and 7000 there were 6 cases, while the highest was 8000. 

The differential counts made in 7 cases were all practically normal 
(even that of the case with a count of 2400 was); s.m. 17.2%, l.m. 2.9%, 
p.m.n. 77.1%, and eos., 1.8%. 

It is interesting that when these 9 cases left the hospital all had gained 
practically the same number of cells, between 900,000 and 1,711,000, 
the mean, 1,100,000. 

The platelets are increased in number as a rule ; in fact, in no condition 
are they as numerous as in chlorosis. They also are large in size. 

The specific gravity of the blood is low, sometimes down to 1.030. This is 
due to the decrease of hemoglobin. In this disease alone does the specific 
gravity of the whole blood run exactly parallel to the hemoglobin content. 
Grawitz states that it varies from 1.035 to 1.045; others say from 1.030 
to 1.050. Grawitz says that if it is under 1.035 some complications must 
be present. 






THE BLOOD 615 

The alkalinity of the blood is normal. 

The isotonicity of the cells is low. 

In the plasma one finds but few changes. There is no blood destruction 
and no hydremia. As the case improves the red cell count rises rapidly to 
normal ; that is, ' ' the anemia is first cured' ' (Graber) , then the light-weight 
cells are slowly replaced by those more normal in size and weight. Yet all 
the variations in the count need not necessarily mean a new formation of 
cells since the first sign of improvement may be an increase of the specific 
gravity and an increased count due to the dissapearance of some of the 
plasma, as is indicated also by the early polyuria and by the disappear- 
ance of edema. Later, the signs of regeneration appear and also the 
gradual elimination of faulty cells. Then the leucocytes may rise to even 
above normal. 

Aplastic Anemia. — Aplastic, hypoplastic, or aregeneratory anemia are 
names applied by Ehrlich to that type of anemia due to decreased blood 
formation. Clinically it is marked by anemia, a pronounced tendency to 
hemorrhage and a rapidly fatal course. The blood shows a marked anemia, 
even 490,000 per cu.mm., while the hemoglobin falls even faster as is shown 
by the continuous drop in the color-index. None of the evidences of bone- 
marrow regeneration are seen, such as megaloblasts, normoblasts, aniso- 
cytes and stippled or basophilic erythrocytes in the circulation, while the 
absence of bile pigment in the skin, urine and blood plasma and the absence 
of hemosiderin in the liver tend to prove that the anemia is due to failure 
in formation and not to abnormal destruction of red blood cells. The 
platelets are few or quite absent. There is progressively developing leuco- 
penia nearly always to 2000 but even to 140 white cells per cu.mm. Every 
type of leucocyte suffers, but the lymphocytes least. The polymorphonu- 
clear cells, including the neutrophiles, eosinophiles and basophiles diminish 
markedly and progressively. 

At necropsy the red marrow is found to be aplastic, showing an in- 
crease of fat and a diminution in megaloblasts and normoblasts. 121 

Musser m collected 59 cases, including 24 of Cabot. 

An aplastic anemia may be due to a toxin which destroys the 
bone-marrow, as benzol, but would seem a possible and logical termination 
of any long-standing anemia and represents the bankruptcy of the 
hematopoietic tissue. 

In 1907 we 123 reported a case of probable aplastic anemia which seemed 
related to the leukemias. The patient, a girl 19 years of age, died after an 
illness of about 1 month with red corpuscles 724,000, hemoglobin 13% and 
leucocytes 1920. Six days before death the leucocytes were 3800 and the 

121 O'Malley and Conrad, Jour. A. M. A., Dec. 6, 19 19, vol. 73, p. 1761. 

122 Arch, of Int. Med., 1914, xiv, p. 275. 

123 Johns Hopkins Hosp. Rep., Mch., 1907, xviii, p. 82. 



616 CLINICAL DIAGNOSIS 

differential count: s.m. 39%, l.m. and tr. 43%- pmn. 14%, eos. 1%, myelo- 
cytes 0.2%, and unidentified cells 2.5%. 

LEUKEMIA 

Barker has defined the leukemic state as one in which there is definite 
proliferation of the leukopoietic tissues, either myeloid or lymphadenoid, 
and the appearance in the blood of immature white blood-cells, usually 
in large numbers, the degree of whose immaturity is more pronounced the 
more acute the cases. 

By this definition of leukemia he excludes Hodgkin's disease and the 
ordinary leukocytoses and lymphocytoses, since in these processes neither 
the blood picture nor the changes in the tissue characteristic of leukemic 
states are present. It also excludes the aleukemic lymphadenoses and the 
aleukemic myeloses (pseudoleukemia) in which, though the histological 
changes in the hemopoietic tissues may be identical with those of leukemia, 
the characteristic blood picture is absent. The aleukemic lymphadenoses 
and the aleukemic myeloses may, however, be closely allied to the leukemic 
states since acute leukemic states are often preceded by aleukemic stages. 
Moreover, in the course of an outspoken leukemia the hematological pic- 
ture may change to that of an aleukemic state. 

From the present point of view of the clinical laboratory, leukemia is a 
disease characterized by the constant presence in considerable numbers in 
the blood of white cells not found normally in the peripheral circulation. 
This is certainly true of the mononuclear granular cells and probably also 
of the non-granular cells present in both forms of leukemia and almost ex- 
clusively in one. These cells are supposed to be immature forms of the 
ordinary leucocytes which because of some disease involving the blood- 
building tissues are extruded into the peripheral circulation. The essence 
of leukemia is, therefore, a marked change in the blood formula. There 
is in leukemia, as a rule, also a great increase in the leucocyte count and 
yet this is not constant while during the periods in which the total count is 
normal the diagnosis can often be made from the presence of these 
abnormal cells alone. 

Leukemia has long been rated among the primary anemias although 
the reasons for this no longer have force. Anemia does, as a rule, develop 
during the course of a leukemia, not as a necessary part of the clinical 
picture, but rather of the cachexia which, sooner or later, always develops. 

According to the blood picture three forms of leukemia may 
be recognized. 

(1) Lymphatic leukemia, lymphadenoid leukemia, "lymphemia," in 
which the increase is of the non-granular cells. 

(2) Splenomyelogenous leukemia, myeloid leukemia, "myelemia," or 
"true leukemia," in which there is an absolute increase of all leucocyte 



THE BLOOD 617 

forms and the presence of mononuclear granular cells never normal in 
the circulation. 

(3) Mixed leukemia, in which the features of both of these 2 forms are 
combined. 

Of all 3 forms, acute cases may occur. 

Leukemia differs from a leucocytosis, not so much because the white 
cell-count usually is much higher, for it often is not, but because the blood 
condition is more permanent and not ephemeral as is a leucocytosis and 
because of the large numbers of abnormal cells (in leucocytosis there may be 
a few present) . 

Splenomyelogenous Leukemia (Plate I). — In splenomyelogenous leu- 
kemia all the granular cells are markedly increased, especially the neutro- 
philes, but also the eosinophiles and basophiles. Abnormal (so far as the 
circulation is concerned) immature forms of each are present together with 
all transitions between these and true leucocytes. The non-granular cells 
are also very much increased, some of which probably are immature 
forms. 

In many cases of this form of leukemia the total volume of blood would 
seem to be increased. This is indicated during life by the dilatation of the 
veins and later by the autopsy. Towards death, however, a diminution in 
the total blood volume would seem to occur. 

Grossly, the fresh blood may look normal even though the leucocytes may 
almost equal in number the red blood-cells. In other cases it has a pale, 
rather opaque appearance and flows sluggishly as though it were thick. 
It is hard to get good smear preparations (which appear granular) ; the diag- 
nosis has often been made in this way, the fresh blood resembling "chocolate 
mixed with cream " (methemoglobinemia?). This must be very rare since 
so many have never seen it. If a larger volume of blood be allowed to settle 
and coagulate, a grayish -white layer will form on the top of the clot 
which may suggest the diagnosis. Coagulation is slow and in the severe 
cases sometimes absent. 

As a rule the red cell-count is diminished (Gr awitz said it always is unless 
some factor is present which concentrates the blood). In none of Taylor's 
cases was it above 4,000,000. The cachexia, slight jaundice, increased 
urinary pigment and the deposit of iron in the various organs show the ac- 
tion of some hemolysin, although the anemia may in part be due to the 
hemorrhages which are so common. Cabot's average count was 3,120,000 
and Osier's, 2,285,000. In 9 Johns Hopkins cases with n admissions, the 
lowest count was 1,640,000, the highest 3,800,000, and the mean 2,800,000. 
It may be almost as low as in pernicious anemia. As the leucocytes increase 
the reds decrease and vice versa. There are exceptions, however, and the 
count may remain almost normal for a long time. This oligocythemia may 
persist during the periods when the leucocyte count is normal and the pa- 
tient feels better. If the patient were seen then for the first time a diagnosis 



618 CLINICAL DIAGNOSIS 

of pernicious anemia would certainly be logical (Taylor). The subjective 
condition of the patients certainly depends very little on the mere count of 
the red cells for they feel well enough to go home when this has changed very 
little from that on admission. 

The anemia is of the chlorotic variety, i. e., the cells are light in weight. 
They show remarkably little degeneration, although endoglobular areas do 
appear. In some severe cases the reds are quite normal. Microcytes and 
macrocytes are rare; a few poikilocytes are found in all cases. The poly- 
chromatophilic degeneration and the basophilic granules are common, yet 
are never marked. Biermer's test was found positive in 2 cases. 

There is no disease in which normoblasts may be found more constantly 
or in greater numbers than in myelogenous leukemia, even in the cases with 
a mild anemia, yet their absence is not against this diagnosis. Megaloblasts, 
and even gigantoblasts 20/1 in diameter, are sometimes found. This megalo- 
blastic feature of the blood while common in leukemia is not as marked as in 
pernicious anemia. Microblasts also are met with. Karyokinetic figures in 
all stages of division may be best studied here. Of Taylor's 16 cases, in 2 
the number of nucleated reds varied from 60,000 to 70,000 per c.mm. and 1 
of the first effects of the arsenic was to reduce their number. It is of interest 
that marked rises in the white count are accompanied by rises in the number 
of nucleated red cells also. 

The hemoglobin is reduced, the color-index being about 0.6. Osier's 
average of hemoglobin was 42%. In 9 recent cases the mean Hb was 30% 
and the mean index, 0.54. The hemoglobin is hard to estimate since' the 
leucocytes render the blood quite opaque. 

From the appearance of the leucocytes of the fresh specimen of blood the 
diagnosis of myelogenous leukemia may sometimes be made at a glance, not 
so much because of the large number of leucocytes as because of the large 
number of immature cells which normally are never present. The count in 
a simple leucocytosis may be as high as that in some cases of leukemia while 
in some cases of leukemia the count is normal. It is the blood formula which 
is important ; it is the large and constant number of abnormal cells. In some 
post-febrile conditions the blood formula may for a short time suggest 
leukemia since it contains a few neutrophile myelocytes but rarely if ever 
does it contain eosinophile myelocytes or an increased number of basophiles. 

Counts of 500,000 leucocytes per 1 c.mm. are not rare. Cabot's average 
on the first visit was 438,000 and Osier's, 298,700. While the counts in any 
given case may vary, they do not as much as might be supposed. The daily 
counts maintain approximately the same level for weeks. We have not 
seen as much daily variation as some have mentioned. We counted the 
blood of several at short intervals. In one case counted each 4 hours the 
counts were 146,000, 134,000, 140,900, and 143,200. More marked varia- 
tions do occur, as in 1 case with 122,500 at 10 A.M. and 235,000 at 4 P.M. 
of the same day. Some cases have quite high counts, over 400,000; but of 



THE BLOOD 619 

the most the counts range from 100,000 to 300,000 (63% of 51 cases), while 
in fewer are they below 1 00,000. The same case on different admissions may 
have very different counts, but during any one stay in the hospital it keeps 
within narrow limits. There are periods when the count is normal, yet even 
then the differential count will usually give the diagnosis. (In 3 of Taylor's 
cases, however, there were no qualitative changes.) 

All of the cells of normal marrow appear in the blood in myelogenous 
leukemia. Among these the neutrophile myelocytes predominate. Some of 
these myelocytes are very large, even 30/* in diameter (Cornil's myelocytes), 
with a large chromatin-poor nucleus, often in an eccentric position, which 
stains so palely that it is hard to make out. These cells are seen only in 
leukemia and in some diseases of children. Other myelocytes are about the 
size of an ordinary leucocyte and have a round nucleus which stains well. 
This is the form seen in the inflammatory leucocytoses. And, finally, one 
usually finds some dwarf myelocytes about the size of a red blood-cell. 
Mitoses of the nuclei of myelocytes are more or less common. The number 
of granules in myelocytes varies considerably ; the protoplasm of some is full, 
others have but a few, while there are some concerning which there is doubt 
whether they are granular or not. Grawitz called attention to large non- 
granular cells with a large pale nucleus (which also may appear free of pro- 
toplasm) which disintegrates rapidly; others are of medium size with baso- 
philic protoplasm which stains intensely and a medium-sized nucleus; in 
other similar cells beginning granulation can be seen. 

Eosinophile myelocytes are present sometimes in large numbers but 
they are never as numerous as is the finely granular form. All transitional 
forms are seen between these and eosinophile leucocytes. 

While the polymorphonuclear neutrophils are relatively diminished 
(Cabot's average was 46%) their absolute number may reach even 50,000. 
Anomalous forms are common: some are very large, even 20/z in diameter, 
while some are small, or dwarfs, 4/1 in diameter. This variation in size 
never appears in a leucocytosis. Again, cells with unusually shaped nuclei 
are found and cells with more than 1 form cf granule. In 1 case all the cells 
were described as non-granular. The plasma is full of free granules from 
many cells which have disintegrated. 

The percentage of lymphocytes is reduced, the average being 10.6%, 
but their absolute number usually is increased. These cells vary much in 
size. Among them are some which can with difficulty be told from myelo- 
cytes. One finds also the large mononuclear cells which are numerous 
enough in the marrow but which never reach the blood normally or in other 
diseases than leukemia. Some have very irregular shapes, some a few 
granules. 

The large lymphocytes have a scanty, ragged protoplasm and a large, 
chromatin-poor nucleus. These are Frankel's unripe cells, supposed by 
some to be characteristic of acute leukemia but which appear also in the 



620 CLINICAL DIAGNOSIS 

chronic types. Large mononuclears, both those of the normal blood and 
those resembling myelocytes, appear in large numbers. Of the latter 
the nucleus is often very basophilic. Their protoplasm is finely fibrillar 
and distinctly basophilic or acidophilic. These cells before Ehrlich's stain 
was used were reckoned as myelocytes. 

Large phagocytes (splenic cells?) are sometimes present and in one 
case numbered 1.2% of the 216,000 leucocytes. 

In this disease there is usually an absolute increase of the coarsely gran- 
ular cells. Ehrlich indeed stated that he would not make the diagnosis of 
leukemia unless more than 250 of these cells per 1 c.mm. were present. 
Since then several cases of leukemia have been reported 124 in whose blood 
at times not a single eosinophile cell could be found and others in which 
these cells fluctuated much in their numbers. As a rule the minimal 
number of eosinophiles in leukemia is about 3000, the average percentage 
5.1 and the average absolute number 11,000. All forms of these cells cor- 
responding to the finely granular cells occur : The large eosinophile myelo- 
cytes (formerly said to be the characteristic cell of leukemia), the medi- 
um-sized and the dwarf eosinophile myelocytes and the ordinary leucocytes. 
The eosinophile myelocytes may in this disease make up the majority of 
the coarsely granular cells. 

Ehrlich considered that leukemia is the one condition in which 
there is an absolute increase of the number of Mastzellen. They may 
even outnumber the eosinophiles. In one of Lazarus' cases they reached 
47%; in one of Cabot's 10%, while Taylor mentions a case with an abso- 
lute count of basophiles of 140,000. Taylor also states that in two cases 
no Mastzellen were found. 

Charcot-Leyden crystals may be found in leukemic blood which 
has stood for a while, but they ma}^ be found also in fresh blood obtained 
by splenic puncture. Some observers, however, have never found them. 
They are normal in the bone-marrow and are present wherever the eosin- 
ophile cells are increased. 

Bezancon and Labbe say that leucin spherules also will separate sponta- 
neously from leukemic blood. 

Ehrlich considered that the diagnosis of myelogenous leukemia could be 
made from the examination of a smear alone provided one found neutrophile 
myelocytes, eosinophile myelocytes, an absolute increase of eosinophiles 
and of Mastzellen, atypical cells, especially the dwarf eosinophiles and 
neutrophiles, cells in mitosis and, lastly, a large number of nucleated reds. 
Yet, for a while at least, any one of these points may fail. 

Many of the leucocytes in leukemic blood show signs of degeneration. 
Ewing considers that eosinophile myelocytes with granules of very unequal 
size and density of stain are pathognomonic of myelocythemia. The pro- 

124 See Simon, Am. Jour. Med. Sci., No. 125, 1903. 



THE BLOOD 621 

toplasm of some cells is swollen, hyaline, or vacuolated. One finds many 
nuclei surrounded by free granules, the protoplasm evidently having dis- 
integrated. Karyolysis, vacuolation and karyorrhexis of the nuclei are 
common; pycnosis perhaps less so. 

The diagnosis of leukemia cannot be made from the total white cell- 
count alone, since the count in pneumonia may rise above 100,000 and that 
in leukemia may drop to normal ; and yet the former high counts are very 
temporary, while those of leukemia continue for a long time. The mere 
presence of myelocytes is not sufficient for diagnosis, since in cases of ex- 
treme leucocytosis a few true myelocytes may be found. These, however, 
are very few in number, are about the size of the ordinary leucocyte and are 
never the very large cells which are seen in leukemia; also in leucocytosis 
the eosinophiles and Mastzellen are not increased. In children especially 
the diagnosis is difficult. Indeed autopsy alone may decide it. 

The white cell-count of 1 case on first admission was 443,000. Fourteen months 
later the patient was readmitted with a count of 9700 and discharged 20 days later with 
one of 100,000. The lowest count during this admission was 6000, of which 3.8% 
were small mononuclears, 3.6% large mononuclears and transitionals, 70.8% 
polymorphonuclear neutrophiles, 3.8% eosinophiles, 8% neutrophile myelocytes 
and 7.6% Mastzellen. There were 2.3 normoblasts, 15 intermediates, and 5 megalo- 
blasts per 1000 leucocytes. Hence even on that day a diagnosis could have been 
made from the blood formula alone. 

With improvement in the condition the count may drop to normal. 
At such times the formula may remain leukemic or may become normal, in 
which case the condition would closely resemble pernicious anemia. And 
indeed cases have been reported as changing to pernicious anemia and vice 
versa. Following the long-continued use of arsenic and of benzol the count 
may drop in a remarkable way usually to rise again after the drug is dis- 
continued. Turk 125 mentions a case in which after arsenic treatment the 
leucocytes, which had ranged from 2 58,000 to 3 70,000, dropped to from 3000 
to 6000 of which 0.5% were myelocytes and 6.6% Mastzellen. It is a ques- 
tion how much improvement these low counts indicate since they may be 
due to "exhaustion" of the bone-marrow. Following X-ray treatment re- 
markable drops have been reported, e. g., from 693,000 to 6300. (In this 
case the leukemic character of the blood was never lost.) But the most 
remarkable falls have followed the use of benzol. In the case reported b} r 
Barry and Ketcham these cells fell from 150,000 to 3000.* 

Radium also has a most definite effect on the blood picture. 126 In cases 
which have received no previous similar treatment the number of white 
cells usually begins to fall in from 24 to 72 hours after the radium is applied. 
The decrease in the leucocytosis is often rapid and continues for days and 
even several weeks after the radium was administered. In one patient the 

125 Deut. med. Wochenschr., 1904, No. 50. 

126 Peabody, Boston Med. and Surg. Jour., Dec, 191 7. 
* Jour, of Ind. State Med. Assoc. Dec, 1916. 



622 CLINICAL DIAGNOSIS 

white count dropped from approximately 100,000 to 6100 in 25 days, ra- 
dium having been given on the first and thirteenth days only. There is a 
change also in the differential count. Myelocytes and immature forms 
of polymorphonuclear leucocytes become less prominent and. a larger 
proportion of adult polymorphonuclear cells is found. Patients with an 
anemia, who respond well to treatment, show a rise in hemoglobin and in 
the red cell-count. On the other hand the development in a case under 
observation of an anemia and the occurrence of many nucleated red cells 
is to be regarded as a serious sign. An important point to bear in mind is 
that the development of anemia may be the result of too much radiation. 
The infectious diseases, especially typhoid fever, influenza, miliary tu- 
berculosis, et al. t have a remarkable effect not only on the blood picture 
but also upon the blood-forming organs of leukemic patients. In Dock's 
case 127 of grippe, the cells fell from 367,000 to 5000 but in 6 weeks had 
returned to 157,000 and in 1 year were 461,000. The fall is sometimes ex- 
treme as from 40,000 to 470. Some of these cases preserve in the low 
counts the leukemic formula, others do not. When the count falls there 
is also temporary reduction in the size of the blood-building organs, 
but not always, as in a case reported by McCrae. Cases are on 
record of leukemic patients who have died of infectious diseases and in which 
at autopsy all signs of leukemia had disappeared from the bone-marrow. 
In other cases, however, the count rises instead of falls. In Miiller's case of 
sepsis the leucocytes dropped from 246,000 to 57,300 and then rose; in v. 
Limbeck's case of pneumonia they fell from 140,000 to 43,500 and then when 
the other lung became involved, rose to 172,000. As the count drops the 
percentage of polymorphonuclears rises and the picture thus approaches 
that of a leucocytosis. 

Late in the disease there may be a marked predominance of large non- 
granular leucocytes. It is possible that some of these are myelocytes with- 
out granulation, the body having lost its power to form the neutrophile 
material (Ehrlich). 

In myelogenous leukemia the count of platelets is markedly increased. 

The water content of the blood is increased to from 81 to 88%. 

The specific gravity of the blood is low, even 1.036; that of the plasma 
is about normal. 

The alkalinity of the blood in leukemia is somewhat decreased by the 
organic acids formed from the breaking down of leucocytes. Formic, acetic, 
lactic and succinic acids have been found in the plasma. The xanthin bodies 
of the plasma are increased. Deuteroalbumoses have been found. These are 
not present in lymphatic leukemia and are supposed to be digestive prod- 
ucts of the leucocytes by a ferment provided by the polymorphonuclear 
cells. Taylor says that the nitrogen of the leucocytes is almost double. 

127 Am. Jour. Med. Sci., 1904, vol. cxxvii. This is a very exhaustive study of this 
subject with a review of 50 cases. 



THE BLOOD 623 

Nucleo-albumin is found in the serum, and 22.6 mgms. of uric acid per 100 
c.c. of blood. The coagulability of the blood is sometimes so increased that 
the red blood-cells cannot be counted with a pipette. 

Lymphatic Leukemia (Lymphemia). (Plate II, A, B, C). — In lymphatic 
leukemia there is a marked increase of the mononuclear non -granular cells. 
Despite the name, these cells are not all typical "lymphocytes" in mor- 
phology, perhaps not in origin, but are mononuclear, non-granular cells of 
many sizes and forms. 

While a variety of these cells may be present 1 particular type usually 
predominates in each case. In most cases they all are small with a very 
narrow ragged rim of protoplasm; in others they all are of the large lym- 
phatic type; in other cases the majority resemble the transparents and in 
still others the transitionals of Uskow. In some cases the protoplasm of 
these large cells seems more basophilic; in others more acidophilic. Some- 
times enormous cells are found. 

In lymphatic leukemia there always is a proliferation of lymphatic 
tissue somewhere in the body. In chronic cases the peripheral lymph- 
glands are enlarged yet in more acute cases none of these glands may be 
palpable. In some cases there are large masses of lymphatic tissue along 
the intestines while in still others the lesion would seem to be limited to the 
bone-marrow. Some interesting cases begin with large peripheral lymph- 
glands and a normal blood picture but later, as the leukemic condition ap- 
pears, these glands diminish in size. One patient in September had normal 
blood and a general glandular enlargement but the following January he was 
admitted with smaller lymph glands and a leucocyte count of 110,000, 
chiefly small cells. 

Pappenheim suggested that the leukemia may not begin until the disease 
of the lymphatic tissue has reached the marrow. 

The anemia is more often marked in lymphatic than in splenomye- 
logenous leukemia and yet the red cell-count may remain normal for some 
time. Later, however, a cachexia is almost inevitable and with it an anemia. 
In two chronic cases the count persisted above 4,000,000 during a long stay 
in the hospital and until death. On the other hand the anemia may be 
extreme as in a remarkable case reported (verbally) by Dr. Hazen, of 18 
months' duration, with red cells 960,000 and leucocytes 250,000, nearly 
all of them small lymphocytes. Cabot's average on the first admission was 
2,730,000; Osier's 2,294,000; that given by Hirz and Labbe is 1,829,000 and 
that by Petit and Weil, 1,292,000. The red cell count is said to be lowest 
in those cases which autopsy shows have most involvement of the bone- 
marrow and also in the more acute cases and in these there always is little 
peripheral glandular enlargement. Nucleated reds are rarely found. Von 
Limbeck describes them as astonishingly scarce. In some very severe 
cases, however, they may be as numerous as in splenomyelogenous leukemia. 
In 1 of our cases in which the total white count was 12,000 there were 15c 



624 CLINICAL DIAGNOSIS 

normoblasts, 169 intermediates and 20 megaloblasts per 1000 leucocytes, 
i. e., a typical megaloblasts crisis. 

The red cell-counts may remain very constant, while those of the leuco- 
cytes are fluctuating widely. In 1 case on 1 day there were 2,640,000 red 
cells and 105,000 leucocytes, 9 days later these counts were 2,750,000 and 
328,800, 2 weeks later 2,892,000 and 410,000, finally in 2 days 2,928,000 
and 480,000. 

In a case with pleural and ascitic effusions (chylous) repeatedly tapped 
and with profuse diarrhea, who died of streptococcus septicemia, the white 
cells showed very slight variations. On admission they numbered 133,400; 
then rose to 242,000 and at death numbered 133,000; the red cells of this 
case numbered 4,912,000 on admission and 5,340,000 at death. Several 
counts were above 5,000,000. 

The hemoglobin is diminished (Osier's average was 37%). 

The leucocyte counts averaged 144,800 (Osier) and 141,000 (Cabot). 
It may be as high as in the myelogenous form, but this is rare. In this form 
also there may be aleukemic periods which may last even 6 months. Just 
before death the count usually rises. One of our cases had a leucopenia 
of even 1900 cells during the 10 weeks before death. 

The mononuclear cells are usually over 90%, sometimes 98%, and in 1 of 
Osier's cases 99%, of the total white-count. A marked feature of the blood 
picture are the degenerated leucocytes. Even 10% and just before death 
even 75% of them may show degenerations of the protoplasm, or pycnosis 
and fragmentation of the nuclei. It is noteworthy that so few of these cells 
in the blood show mitotic figures since in the bone-marrow their proliferation 
is very active. Grawitz classifies the cases as: those in which the small 
mononuclears, predominate (many of these are very small, even smaller 
than red blood-cells, their protoplasm intensely basophilic and scanty, the 
margin ragged and degenerated, their nuclei round or indented and even 
fragmented with sharp margins and often containing clear areas) ; those 
with a predominance of medium-sized cells with basophilic homogenous 
protoplasm ; and those in which the cells which predominate are very large 
and are, for the most part, degenerated. Yet all these forms may occur 
together in the same case, and vary in their relative percentage at 
different times. Roser thinks that in those cases in which the lymph- 
glands are particularly involved it is the smaller cells which are increased 
( e - g-> 99% of 117,000), and that in those in which the lesion is particularly 
of the bone-marrow, the larger cells. Grawitz mentions a case in which 
the percentage of the larger cells increased simultaneously with a de- 
crease in the size of the lymph-glands. Wolff thinks we should separate 
lymphatic from lymphoid leukemia, the former being of lymph-gland 
origin, the latter myelogenous. 

Polymorphonuclear granulated cells are rare in the circulating blood. 
Eosinophile cells are usually absent. In a pure case no myelocytes are pres- 



THE BLOOD 625 

ent, although it would be hardly wise to call the case mixed leukemia if 
one were found . Mastzellen are as a rule absent. In this form of leukemia 
an acute infection may cause a drop of the total count or a true leu- 
cocytosis. On autopsy on cases of lymphatic leukemia who died of 
acute infection no leukemic lesions have been found. 

During the periods when the count is low the mononuclear cells may be 
even 90% of the total count. In Wende 's case 128 complicated by a strepto- 
coccus infection the white cells dropped from 45,000 to 1600, but the per- 
centage of small mononuclears only from 95.3 to 88%. As the result of 
an acute infection a few myelocytes may appear. In other cases, however, 
an infection causes a marked increase in the count. In Muller's cases, 
complicated by chronic septicemia, e.g., it rose from 180,000 to 400,000. 

A man was admitted to this clinic with double tertian malaria and a lymphatic 
leukemia of 105,000 cells (small monos. 83.6%; large monos. and tr. 7%; pm. n. 5.8%; 
eosinoph. 0.2%). One week after the malaria was cured the count rose to 328,000 and 
2 weeks later to 480,000 with 97.2% small mononuclears. At this time there were 3 
normoblasts, 3 intermediates and 4 megaloblasts per 1000 leucocytes. 

Von Limbeck considers that the blood picture alone is not enough for 
diagnosis of lymphatic leukemia since in some cases of sarcoma the blood 
presents a similar picture. 

There is a good reason for separating the acute leukemic states from the 
chronic leukemic states, for the mode of onset, the clinical symptoms, the 
course, the duration, the blood picture and at autopsy the histological 
changes, differ. Whether the etiological agent is the same for the 2 groups of 
cases is not certain (Barker) . It is possible that they are identical though 
many investigators believe that the etiological agents are entirely different 
from one another. In favor of the identity of etiology is the fact that a 
chronic leukemia may have an acute onset or an acute termination. 

Acute Leukemia. — Acute leukemia is a form of leukemia characterized 
by its brief course ,— from 6 days to 9 weeks (leukemia acuta et acutissima), 
— the severity of the symptoms, frequency of the hemorrhagic diathesis, 
the rapidly developing cachexia and the rapid death. It occurs chiefly in 
young persons. The great majority of the cases are of the lymphatic type, 
''acute lymphadenoid leukemia," but a few of the myelogenous variety, 
"acute myeloid leukemia," have recently been reported while other cases 
are best described as mixed. 

Cases of the acute myelogenous forms are collected by Gardinier 129 who 
reports 1 and reviews 11 others. 130 In all of these cases the anemia is 
extreme, the red blood-count, even below 1,000,000 cells. In Arneth's case 
the red cell-count was 256,000 per c.mm. and the hemoglobin 10%. 

128 Johns Hopkins Hosp. Bull., October, 1904 

129 See also Billings and Capp, Am. Jour. Med. Sci., 1903. 

130 Am. Jour. Med. Sci., vol. cxxii, 1901. 
40 



626 CLINICAL DIAGNOSIS 

The cases of acute lymphatic leukemia are well reviewed by Rosen- 
berger. m 

Acute leukemia in children is reviewed by Churchill, 132 who reports i 
case and reviews 28 others. The disease occurs even in the new-born child. 
The lowest red count was 750,000 (after a severe hemorrhage). The 
leucocytes varied from 6000 to 810,000 (in a 20-month-old child). The 
counts were always lowest just before death, which a falling count portends. 
Of these 29 acute cases in children, 25 were lymphatic (2 of the small-celled 
type, 3 large, 1 mixed), 1 was myelogenous, 2 were mixed and 1 was uncer- 
tain. In Churchill's case 99% of the white cells were small mononuclears 
many of which were degenerated. The anemia is profound. It is of interest 
that the more acute the case the less evidence is there of involvement of 
lymph-glands and spleen. 

A good illustration of this type is Pfannkuch's case, which ended fatally in 3 days. 
The reds numbered 2,500,000 and the leucocytes 1,000,000 (s.m. 76.5%; neutrophilic 
myelocytes, 10.6%; neutrophile leucocytes, 12.2%). In Turk's case, a good illustration 
of the myelogenous form, the red cell count was 1,060,000; hemoglobin 19% and the 
leucocytes 42,000 (s.m. 14%; pmn. n. 32% and myelocytes 47%). 

The blood picture in a variety of conditions may closely resemble acute 
myelogenous leukemia. These are: an acute exacerbation of a chronic 
myelogenous leukemia; a lymphatic leukemia complicated by an acute 
infection; an acute lymphatic leukemia of the large-celled variety (since it 
is not easy to distinguish these cells from myelocytes) ; an acute infection 
causing grave anemia, in which case even 14% of the leucocytes may be 
myelocytes; an acute exacerbation of pernicious anemia; and, finally, 
malignant disease of bone-marrow. A very large number of nucleated reds 
would suggest this last condition (Billings and Capps). 

In many cases the clinical picture of acute leukemia is that of an acute 
infection and very likely in the future such cases will not be grouped under 
the leukemias. 

There is no type of cell characteristic of acute leukemia: Frankel's 
unripe cells are common, but are found by no means exclusively here. In 
3 of McCrae's 5 cases the small lymphocytes predominated. The cells in 
any given acute case are likely to be much more uniform than in a chronic 
case, yet this is by no means always true. (Plate II, C.) 

The red cells show few changes. As a rule nucleated reds are scarce, 
yet in Herrick's case they numbered 1,800 per cubic millimeter, of which 
some were megaloblasts (but see McCrae's case). The falling count, 
showing rapid blood destruction, may be a striking feature. 

In other cases the leucocyte count is not even above normal although 
the percentage of lymphocytes is quite high (Klein). Such cases may at 
first resemble pernicious anemia. 

131 Am. Jour. Med. Sci., 1904, vol. cxxviii, p. 583. 

132 Ibid., 1904, vol. cxxviii. 



THE BLOOD 627 

The 5 cases of acute lymphatic leukemia in the Johns Hopkins clinic have been 
reported by McCrae. 133 Their duration, which varied from 12 days to 8 weeks, averaged 
6 weeks. On admission the hemoglobin averaged 35.4%; the reds, 1,822,000 and the 
leucocytes, 104,000. The highest white count was 326,000 (hemoglobin 45%; reds 
3,000,000); the lowest 57,800 (reds 748,000; hemoglobin 16%). In 1 case in 15 days 
the reds fell from 3,000,000 to 1,450,000. The color-index is high, from 0.93 to 1.4. 
The small mononuclears varied from 94.2 to 99.4% and in 3 the small lymphocytes 
were the prevailing cell, unusual in this form of leukemia. In 1 case actively motile, 
large lymphocytes were seen. Nucleated reds were absent in 2 cases, were present (2 per 
1000 leucocytes) in 3 and in the fifth case there were 310 per 1000 leucocytes (i.e., 3720 
per c.mm. of which 7% were megaloblasts and 48% intermediates). McCrae empha- 
sizes the high color-index of these and of the cases in literature and finds that of 45 cases 
in 24 the red count was below 1,500,000 and in 38 of the 45 it was below 2,500,000. Of 
40 cases from literature, in 20 the color-index was 1 or above 1. The low red count and 
the high index of the primary anemias seem to him a special feature of this type of leu- 
kemia. A most remarkable case was admitted with 752,000 red cells, hemoglobin 17% 
and 880,000 leucocytes of the large-cell type. 

In 13 cases of acute leukemia collected by McCrae, 134 the anemia was 
severe (the highest red count was 2,350,000; the lowest, 1,000,000) and the 
color-index high. The red cells were of normal appearance. No nucleated 
reds were found in 7 cases, a few in 4 and megaloblasts in but 1. The leu- 
cocytes varied from 21,000 to 209,000. The absolute number of polymor- 
phonuclears was about normal in all cases. In these cases the anemia was 
the important feature. The leukemia would not have been suspected 
without blood examination. 

The cause of leukemia is still to be discovered. Auer 135 described very interesting 
rods in the cytoplasm of the large mononuclear leucocytes of a case of acute leukemia. 
Mention may be made of Lowit's organism (Fig. 134). In cases of splenomyelogenous 
leukemia he described bodies in the large and small mononuclears, both non-granular 
and granular, rarely in the polymorphonuclear cells, never in the red cells and sometimes 
free in the plasma, which he named Hemameba leukemic magna. Their size varies 
much. Some of the groups of such bodies he considers evidence of multiplication. They 
can be stained by a particular method. Lowit found in the blood-building organs navicu- 
lar and crescent-shaped bodies which suggested the coccidia and the hemosporidia and 
which first suggested to him the possible parasitic nature of the disease. He did not find 
them in other conditions, and denied that they are artifacts. Finally he states that he 
got positive results from rabbit inoculation. 

In 1 of 5 cases of lymphatic leukemia he found Hemameba leukemiae parva, which 
was smaller than the preceding and apparently more ameboid. These he found especi- 
ally in the blood-building organs, rarely in the peripheral blood. 

Turk considered these parasites of Lowit to be much altered basophile granules or 
artifacts. By unanimous consent the question has been allowed to drop and yet if all 
these bodies which we have seen in the blood specimens stained with his method are 
artifacts they certainly were beautiful pseudoparasites. 

The frequent association of acute lymphatic leukemia with tumors of the 
thymus has been emphasized by Major 136 and Asbury.* 

133 Brit. Med. Jour., February 25, 1905. 

134 Johns Hopkins Hosp. Bull., May, 1900. 

135 Am. J. Med. Sci., June, 1906. 

136 Johns Hopkins Hosp. Bull, Sept., 1918, xxix, 331. 
* Jour, of Ind. State Med. Assoc. Dec. 1920. 



628 CLINICAL DIAGNOSIS 

Mixed Leukemia. — The cases of mixed leukemia may best be described 
as a lymphatic leukemia with a considerable number of myelocytes, both 
eosinophile and neutrophile, also present.. Since a few myelocytes may 
occur in lymphatic leukemia the term should be used with caution. 

Pseudoleukemia. — Under the heading pseudoleukemia have been 
grouped a great variety of diseases which suggest leukemia clinically but not 
hematologically. Of first importance under this heading are the early stages 
of lymphatic leukemia, before the blood changes appear, and cases of leu- 
kemia during the aleukemic periods. But the group includes Hodgkin's 
disease (which we believe can be excluded by removing a superficial gland), 
tuberculosis of the lymph-glands, lympho-sarcoma and malignant lympho- 
mata (which the pathologists say can be recognized anatomically). Others 
include splenic anemia. It is, therefore, the opinion of some that the exist- 
ence of a "true" pseudoleukemia is still to be proved (Reed) . 

Hodgkin's Disease. — The blood features in Hodgkin's disease are 
those of cachexia. At the onset the count may be practically normal and 
remain so for months despite the rapid growth of the lymph-glands. Then 
slowly develops an anemia of secondary type, often extreme, with at the 
end a count as low as 1,522,000 with degeneration of the red cells, nu- 
cleated reds and poikilocytes (which are noticeably rare except at the late 
stages). There is a slight leucocytosis averaging about 12,000. In the 8 
cases reported by Reed the red count on admission varied from 3,232,000 
to 5,264,000. In 1 case it was 2,670,000 but afterwards improved. These 
cases were, therefore, somewhat anemic, while 2 showed a severe anemia of 
the secondary type. 

In 2 of the 8 cases the small mononuclears were absolutely increased but 
the lymphocytosis of Pinkus is by no means constant. In 1 case the count 
of small mononuclears was 5304 per c.mm. (38.6%) and the next highest 
was 4600 (36.8%). In 2 cases the small mononuclear count was low; in 
one 310 (2%) and in another 940 (9.4%). Grawitz considers that the 
differential count is of no aid in the diagnosis of Hodgkin's disease but 
is in the prognosis since a slight increase indicates improvement and a de- 
crease, the reverse. 

Tuberculosis of the Lymph-Glands. — In cases of general glandular 
tuberculosis there may, for a time, be a normal red and leucocyte count but 
one more often finds a secondary anemia accompanying the cachexia. Some 
of the lowest white counts of all have been reported in this condition, 
e. g., 300 leucocytes per cubic millimeter (Futcher). 

We studied 12 cases of this disease in the Johns Hopkins clinic. Four had a leuco- 
cytosis of from 11,000 to 29,000 and a slight secondary anemia. In 2 cases the red cell 
counts were 3,600,000 and 3,700,000; and in 6 cases it varied from 4,000,000 to 5,000,000; 
the hemoglobin was so much reduced that in 6 cases the index was below 0.6. In this 
disease a leucocytosis as a rule means a secondary infection. 

A case like the following is a puzzle for diagnosis. The woman, aged 50 years, was 



THE BLOOD 629 

first admitted with a red count of 4,000,000, hemoglobin 50% and leucocytes 8000. She 
had tuberculosis of the lungs and swollen lymph-glands, 2 of which 'had been removed 
with an interval of 1 year between operations and both pronounced tuberculous. She 
had night-sweats and lost weight. Three months after the above blood-count the glands 
began to swell enormously. The red cells then were 3,000,000 and the leucocytes 80,000, 
96% of which were polymorphonuclear neutrophiles. A little later the count rose to 
120,000. The lymph-glands and spleen became enormous. She received X-ray treat- 
ment and in 3 weeks the leucocytes fell to 16,000 and the reds to 2,100,000. She died 
soon after. 

Leukanemia is the name given by v. Leube to a group of cases which have 
the features of both acute leukemia and of pernicious anemia. In the blood 
of cases with a severe anemia appear nucleated reds of all varieties and a 
normal or increased white count. In some of the cases many myelocytes 
appear present but no eosinophiles while other cases resemble lymphatic 
leukemia. The anemia usually precedes the increase in the count of the 
white cells. This blood picture is met with in a variety of conditions 137 
including injuries, hemorrhages, intoxications, infections, malaria, malig- 
nant growths, etc. 

BLOOD IN ACUTE DISEASES 

Malaria. — The presence of an anemia is important in the diagnosis of 
malaria since it is one of the earliest symptoms. In an acute case the count 
of red cells may, within a few days, drop from 5,000,000 to even 500,000. 
This is the result both of the direct destruction of the corpuscles by the 
intracellular parasites and of the action of a toxin which can produce 
rapid hemolysis. Such cases are, however, very rare. In the average case 
of tertian and quartan malaria the red cell count decreases slightly after 
each paroxysm but in the asstivo-autumnal fever with chills there may be a 
drop of 1,000,000 cells after a single paroxysm and a further fall between 
the chills. 

In 54 cases of asstivo-autumnal malaria the red cells numbered between 1,000,000 
and 2,000,000 in 2 cases; between 2,000,000 and 3,000,000 in 12; between 3,000,000 
and 4,000,000 in 20; between 4,000,000 and 5,000,000 in 12; and above 5,000,000 in 8. 
In 56 cases of tertian malaria the figures for these same limits were 1, 10, 28, 13, and 
4 respectively. The mean count for each was 3,500,000. It is seen that in the Balti- 
more cases at least these 2 forms differ but little. In this climate the pernicious cases 
are rare. 

In Grass's case there was a loss of 4,000,000 cells in 6 days. In one case 
the count at the end of 30 days was 5,000,000. The greatest fall in the count 
follows the earliest paroxysms, less later, until finally the count remains 
almost stationary despite repeated paroxysms. In cases with pernicious 
malaria and hemoglobinemia the anemia becomes grave and poikilocytes, 
endoglobular degenerations, occasional shadows, fairly numerous nucleated 
reds, increased platelets and a leucocytosis appear. 

137 Luce, Deut. Archiv. f. klin. Med., 1900, vol. lxxvii, p. 215. 



630 CLINICAL DIAGNOSIS 

The regeneration of the cells in tertian and quartan malaria is rapid 
and anemia develops only in long-standing cases. In asstivo-autumnal 
malaria the recovery is slower, the new cells are pale and abnormal in size 
and shape, nucleated reds are common, regeneration is slow and a grave 
anemia may result. This slowness in regeneration is due in part to the ex- 
tensive necrosis and resulting fibrosis of the bone-marrow, which may be the 
chief seat of the infection, and to the accumulation of pigment in this tissue. 

The leucocyte count in malaria is almost always below the normal ex- 
cept in the grave pernicious forms. It rises slightly (to 6700, some say to a 
true leucocytosis) just before the paroxysm, and then falls steadily even to 
2000 or even 1000 cells (average 2300) reaching a minimum at the time 
the temperature is subnormal. 

In 82 recent cases of aestivo-autumnal malaria the leucocyte counts were: from 
1000 to 2000, 3; 2000 to 3000, 8; 3000 to 4000, 21; 4000 to 5000, 15; 5000 to 6000, 14; 
6000 to 700G 8; 7000 to 8000, 4; 8000 to 9000, 2; 9000 to 10,000, 2; above 10,000, 5. 
(In 1 of these, a pernicious case, the count was 14,500. The mean count was 3500.) 

In 70 cases of tertian malaria the figures for these same limits were 2, 5, 11, 18, 10, 
10, 5, 2 and 2; above 10,000, 5. The highest count was 16,500 and the mean 4500. 

The differential count in cases of malaria shows a relative decrease of the 
polymorphonuclear neutrophiles and an absolute increase of the endothelial 
leucocytes. The mean averages found by Thayer are: s.m. 16.9%; l.m. 
16.9%; pmn.n. 65%; eos. 0.9% and in grave cases 2 to 3% of myelocytes 
(Cabot) . The increase of the large mononuclears (endothelial cells) is very 
pronounced in the apyre tic periods and usually absent in the pyretic periods. 
If in a case of intermittent fever these cells do not increase as the temper- 
ature falls the evidence is against malaria. The high percentage of these 
cells is very valuable in the diagnosis of cases which have been taking 
quinine and therefore have no parasites in the peripheral blood. Stevens 
and Christophers say that in the Tropics a blood formula with 1 5% or more 
of large mononuclears indicates malaria, while with 20% or more one can 
almost always find the parasite. This group of endothelial cells, which vary 
in size from lymphocytes to the largest cells of the blood, are slightly 
ameboid and distinctly phagocytic, in fact are the chief phagocytes in 
malaria. Many of them contain pigment granules. Such cells are almost 
as valuable in diagnosis as is the parasite itself. 

Pigmented endothelial leucocytes may be found in the peripheral cir- 
culation in cases of aestivo-autumnal malaria most of the time but in tertian 
and quartan malaria only immediately after a paroxysm. They are said 
to become necrotic rapidly and so to disappear soon from the circulation. 

Stevens and Christophers cite Bastianelli's fatal comatose case of 
aestivo-autumnal malaria with s.m. 19.1%, l.m. 41%, pmn.n. 39%, and 
eos. 0.06%; also Panse's case with a temperature of 37.2°C, and s.m. 
18.1%, l.m. 26.4%, pmn.n. 55.3%; and another case with temperature 
normal, with s.m. 14.8%, l.m. 46.7%, and pmn.n. 38.5%. 



THE BLOOD 631 

In i of the Johns Hopkins hospital cases of tertian malaria the leucocyte count was 
16,500 of which 38.3% were large mononuclears. In a case of aestivo-autumnal malaria 
the leucocyte count was 6000 of which 26% were large mononuclears. In another case 
with 4000 white cells, 22% were endothelial cells. 

A leucocytosis is rarely met with in malaria, except in the pernicious 
forms. In one case in that clinic the count one hour before death was 
50,000 of which the large mononuclears and transitionals were 18%, 
and the polymorphonuclear neutrophiles, 58%. A polymorphonuclear 
neutrophile leucocytosis may develop with an attack of malarial hemo- 
globinemia and persist for some time. Those seen during the death 
agony are due to complications. A definite leucocytosis, 'sometimes with 
increased eosinophiles and with myelocytes, is common with the post- 
malarial anemias. 

Bignami and Dionisi classified the anemias of malaria as follows: (1) 
The secondary anemia of the chlorotic type which follows acute malarial 
fever in which there are a few nucleated reds, leukopenia and an increase 
of the large mononuclear leucocytes; (2) cases resembling primary per- 
nicious anemia, usually fatal, with extreme oligocythemia, marked 
poikilocytosis, high color-index, nucleated reds usually megaloblasts, 
leucopenia and lymphocytes relatively increased; (3) rapidly fatal cases 
without any signs of regeneration which may have started like a simple 
secondary anemia. This anemia is very similar to that which follows a 
severe hemorrhage. (4) Chronic, grave, secondary anemias of the chlo- 
rotic type, without nucleated reds and with leucocytes much reduced. This 
is seen in chronic malarial cachexia, and is due in part to degenerative 
changes with sclerosis and pigmentation of the bone-marrow. (Thayer). 

Septicemia. — That septicemia does not always cause a leucocytosis is 
seen in typhoid fever and acute miliary tuberculosis, but those due to 
pyogenic organisms usually do. In the streptococcus and staphylococcus 
septicemias, e. g., the puerperal infections, the anemia develops more rapidly 
than in any other acute disease. The loss usually is from 200,000 to 1 ,000,000 
cells per week but Grawitz mentions a case of acute streptococcus 
septicemia with hemorrhages in which the red cells fell in 24 hours 
from about normal to 300,000. The qualitative changes of the blood are 
marked; degenerations, poikilocytosis, polychromatophilia, etc. Nucleated 
reds seldom are present in large numbers. The leucocytes vary as the 
patient's resistance. In some cases the counts are very high, in other cases 
even subnormal. 

We have had 26 cases of well-marked septicemia. The drop in the count between 
admission and death was from 900,000 to 1,600,000 cells. The final red counts in 15 
fatal cases were: from 1,000,000 to 2,000,000, 2; from 2,000,000 to 3,000,000, 3; from. 
3,000,000 to 4,000,000, 4; and above 4,000,000. 6. 



632 CLINICAL DIAGNOSIS 

It is thus seen that there are 2 groups of cases, those with high and 
those with low counts, and that the infection is not serious because of 
the anemia alone. 

In 4 cases there was no leucocytosis at death. In 21 cases the leucocytes varied 
from 11,000 to 47,000. These cases showed great variations in the blood formula. In 
1 case with 8000 leucocytes 96.6% and in another with 10,400 leucocytes 92.6% were 
polymorphonuclears. In a case of gonorrheal septicemia the red count at death was 
2,318,000, hemoglobin 30% and the leucocytes 47,000. 

Chronic Septicemia of cryptic origin often would pass unrecognized were 
it not for the anemia. Cases of chronic abscess formation may have but 
little anemia. In Ewing's case of empyema the red blood cells, after a 
duration of 1 year, numbered 1,800,00a and the hemoglobin 25%. In cases 
of pelvic abscess of even 2 years' duration only a slight anemia may develop. 

Blood in Endocarditis. — In simple (rheumatic) endocarditis the blood 
cultures are sterile in 90% of the cases (some would exclude from this group 
all in which the cultures are not sterile) and cultures from the vegetations 
made at autopsy show no constant organism. 138 

Instead of the terms "ulcerative," "malignant," or "infectious" 
endocarditis, Libman 139 suggests the term "bacterial" endocarditis and if 
the causal organism is known he substitutes the name of this for the 
word "bacterial" (e. g., acute or subacute streptococcus endocarditis, etc). 

In all cases of septicemia we may expect to find the heart valves in- 
volved . Especially is this true of streptococcus, pneumococcus, gonococcus, 
meningococcus and influenza septicemias. 

Subacute streptococcus endocarditis, a condition usually fatal, is a 
disease of definite bacteriology due to a constant individual type of strep- 
tococcus which belongs to the saprophytic group, is of low virulence, is 
non-hemolytic and may (viridens) or may not (saprophytics) be green 
producers. Biologic and immunologic tests, however, fail to show any 
constant identity between the individual streptococci concerned in pro- 
ducing this disease. 140 Libman especially has described this type under the 
name subacute bacterial endocarditis due to streptococcus viridens. 
Swift and Kinsella (ibid., p. 381) warn us not to include in this group all 
cases with endocarditis from whose blood viridens may be cultivated since 
other characteristic signs as petechiae, embolic lesions and a progressive 
downward course are also necessary for that diagnosis. 

Typhus Fever. — We have studied the records of but 4 cases of 
typhus fever : 

Case I. — Male of 36 years. On admission: red cells 5,400,000, hemoglobin 72% 
and leucocytes 18,600. The temperature varied from 103° to 104 F. On the eighth 

138 Swift and Kinsella, Arch, of Int. Med., Mch., 1917, xix, p. 381. 

139 Am. Jour. Med. Sc, 1912, vol. 144, p. 313. 

140 Kinsella, Arch, of Int. Med., Mch., 1917, xix, p. 367. 



THE BLOOD 633 

day after admission the white count was 25,400; the temperature had then begun to fall. 
Five days later, the temperature then normal, the total count was 24,300; s.m.3.2%, 
l.m. and tr. 6.6%, pmn. n. 90% and eos. 0.2%. 

Case II. — Male of 19 years. On admission: red cells 4,500,000, hemoglobin 70% 
and leucocytes 8600. This count remained normal for 4 days during which time the 
temperature varied from 102 to 105 F. On the fifth day the count was 12,500 and on 
the tenth day, with temperature normal, the total count was 10,000, s.m. 6%, l.m. and 
tr. 4.2%, pmn. n. 89.4% and eos. 0.2%. . 

Case III. — Male of 30 years. On admission: the red cells 5,500,000, hemoglobin 
85%, and leucocytes 7000. They remained normal 3 days during which time the tempera- 
ture ranged from 101 to 103 F. On the fifth day, the day of death, with temperature 
98 F., this count was 38,000, s.m. 5.8%, l.m. and tr. 1.2% and pmn. n. 95%. 

Case IV. — Male of 22 years. On admission: red cells 5,400,000, hemoglobin 85% 
and leucocytes 9200. This count remained normal for 3 days during which the tempera- 
ture varied from 102 to 104 F. On the tenth day the leucocyte count had risen to 1 1 ,600 
but the temperature had already been normal 3 days. On the twelfth day the leucocyte 
count was normal. On the ninth day the leucocyte count was 10,800, s.m. 15%, l.m. 
and tr. 11.5%, pmn. n. 72%, eos. 1.0 and Mastzellen 0.8%. 

From these 4 cases (daily counts were made in all) it is seen that the leucocytes are 
low, even normal, on admission, then when the temperature has begun to fall or is 
already normal they rise to a maximum and then they fall to normal. (Compare with 
influenza and variola.) 

Ewing and Thomas report an absence of leucocytosis in typhus fever. 

Measles and German measles have almost no influence on the red 
blood-cells and cause no, or only a slight, leucocytosis but more often a 
leucopenia (see page 521). In the post-febrile stage the large mononuclears 
are increased. 

Plantenga found in the 13 cases of measles and the 9 of Rotheln which 
he studied a neutrophile hyper leucocytosis of even 20,000 cells during the 
prodromal stage. During the eruptive stage this rapidly gave place to a 
hypoleucocytosis, due to the disappearance of the neutrophile cells, with 
sometimes a lymphocytosis and the disappearance of eosinophiles. 

Renaud found in 6 cases that this preliminary leucocytosis reached its 
maximum about 6 days before the rash appeared. This permits one to iso- 
late a suspected case early. 

Tileston could not confirm this leucocytosis during the prodromal stage, 
and thought that all leucocytoses could be attributed to complications. 

We have very little material, but of 9 recent cases in only 1 was the 
count above 8600 during the height of the fever (17,200). 

Scarlet fever causes a slight anemia, the count averaging 4,500,000 
(Reckzeh), and a leucocytosis which develops even 6 days before the rash 
appears and continues for even 12 days into the convalescence after the 
temperature has reached normal. Scarlet fever furnishes an interesting 
exception to the general rule that the leucocyte count runs roughly parallel 
to the temperature. The leucocyte counts vary from about 10,000 to 40,000 
(in mild cases, 10,000 to 20,000; in moderate, 20,000 to 30,000, and in severe 
30,000 to 40,000) according to the severity of the case and its duration. 



634 CLINICAL DIAGNOSIS 

The neutrophile cells are relatively increased (to from 85 to 98%, highest 
in the fatal cases). There is often an early eosinophilia, which is important 
in the diagnosis since it excludes various septic conditions, which reaches the 
maximum 2 or 3 days after the rash appears and disappears with the leuco- 
cytosis. (In other diseases these cells disappear during the fever and return 
with improvement.) Their failure to reappear is considered a bad sign. 

Diphtheria. — In diphtheria a moderate anemia develops which amounts 
to a loss of about 2,000,000 cells at the time of defervescence. During the 
height of the disease, however, the red cell count and the specific gravity 
of the blood often rise. (The injection of these bacilli or their toxins into 
the circulation of animals has a lymphagogue action which results in a 
hypercythemia.) This hyper cythemia occurs most commonly in this of all 
the acute infections. In Cutter's case the cells varied from 7,200,000 to 
7,800,000; in Morse's from 5,000,000 to 5,500,000 during the first, and 
reached 6,800,000 during the second week. With the drop in the count 
nucleated reds and polychrcmatophilic cells appear. A slight leucocytosis 
of from 10,000 to 15,000, due to an increase of the polymorphonuclear neu- 
trophile cells, is the rule but in severe cases the count may reach 17,000 and 
with complications even 30,000. In some fatal cases there is leucopenia. 
Myelocytes, even from 3 to 16%, are often found especially in the fatal 
cases. Morse says: "The examination of the blood in diphtheria is of no 
practical clinical importance in diagnosis, prognosis, or treatment." 

In ordinary follicular tonsillitis the counts are often as high as 
in diphtheria. 

Smallpox. — "No other disease is so destructive to the red blood-cells." 
(Hayem) . A low red cell count is the rule as the temperature falls and yet 
this may be due in part to a dilution of the blood plasma resulting from 
the relaxed vasomotor tone which obtains then. Pick and Weil say that 
there is anemia in the severe, but none in the mild, cases. During the pus- 
tular stage of severe cases, however, there may be a true loss of 2,000,000 
cells while in the hemorrhagic type the anemia is severe. Regeneration is 
slow, lasting about 14 days. Nucleated reds (normoblasts) are rare, ex- 
cept in the hemorrhagic cases in which they may be very numerous. 

From the onset the total leucocyte count is normal but the blood formula 
is very characteristic in this disease. The polymorphonuclear neutrophiles 
are decreased, averaging about 40%, but they may sink to 20 or even to 
14%. The small mononuclears vary from 30 to 40%, the large mononu- 
clears from 4 to 10% and the myelocytes and irritation forms each 2 to 10%. 

Smallpox itself causes no leucocytosis, and yet during the pustular stage 
a leucocytosis is often present. 

Tuberculosis. — Tuberculosis is a disease which is accompanied by the 
greatest variety of blood pictures. In a few cases there is anemia of the 
highest grade (e. g., v. Limbeck's case of tuberculosis of the peritoneum and 
abdominal organs, with a red blood-count of 730,000 and hemoglobin 25%; 



THE BLOOD 635 

such cases are so rare that this one is doubted by Cabot) ; in others, one 
of moderate grade, and often more apparent than real, while in others 
there is none. 

In tuberculosis in general a mild grade of chlorotic anemia is the rule. 
This " pseudochlorosis tuberculosa " occurs in cases with slight involvement 
of the apex (" anemia of onset ") without fever and in tuberculosis of bones 
and lymph-glands. The count is almost normal, the leucocytes normal and 
the hemoglobin somewhat reduced. In other cases there is a lymphocytosis, 
absolute or relative, while in some few cases there is a reduction of the count 
as well as of the hemoglobin. Qualitatively some of the red blood-cells 
(not the majority, as in chlorosis) are rather pale and small. Poikilocytes 
while usually few may be numerous, but not as numerous as in other 
cachexias of the same degree. Maragliano's endoglobular degenerations are 
seen in severe cases, especially in those with mixed infections. Nucleated 
reds are rarely present even after a severe hemorrhage causing extreme 
anemia. This helps in the differential diagnosis between tuberculosis 
and carcinoma. 

Cabot who believes that the tuberculous virus has but little effect on 
the blood thinks that the above-mentioned changes are due to secondary 
infections or to drains upon the proteid of the blood from diarrhea, effusions, 
starvation, prolonged suppuration, etc., and Miller, Lupton and Brown 141 
report remarkably few blood changes in cases of pulmonary tuberculosis 
undergoing sanitarium and tuberculin treatment. 

In tuberculosis without secondary infection, with the possible exception 
of meningitis, the leucocytes are not affected. This is important in the 
diagnosis of tuberculous peritonitis, osteitis and acute miliary tuberculosis. 
There also are no important qualitative changes, for the lymphocytosis with 
a normal count, so often mentioned, is common to all cachexia-producing 
conditions. If a leucocytosis does develop it is of the ordinary inflammatory 
type. The eosinophile cells are increased in some cases with cavity for- 
mation. Since a similar increase follows the injection of tuberculin some 
think it is due to auto-intoxication from the cavity. Myelocytes appear in 
the peripheral blood of advanced cases. 

Chronic Pulmonary Tuberculosis. — Grawitz has divided cases of chronic 
pulmonary tuberculosis into three groups: Group i, with slight in- 
volvement of the apex and without fever. This is accompanied by the 
pseudochlorosis tuberculosa (see above) and a normal leucocyte count. 
Some early cases have practically normal blood. Group 2, of cases of 
chronic phthisis with cavity formation but without other complication and 
with slight temperature, is noteworthy since the blood picture is practically 
normal as regards count, hemoglobin, specific gravity and dry constituents. 
This is remarkable since during the cavity formation there is general 

141 Am. J. of Med. Sc, M^y, 1912, No. 5, cxiii, p. 683. 



636 CLINICAL DIAGNOSIS 

emaciation. The leucocytes are normal or slightly increased, from 10,000 
to 15,000 per c.mm. These patients earlier may have had a distinct 
chlorosis. Group 3 includes cases with hectic fever (supposed by many to 
be due to a secondary infection but by others to be due to a pure infection 
with the tubercle bacillus) in which often a true anemia develops, with 
evidence of blood destruction, which may progress until death. The drop 
of the count in these cases may be very rapid. 

In a recent case of pulmonary tuberculosis 2 days before death the red count was 
1,473,000, the hemoglobin 15% and the leucocyte count 9000 (pmn. n. 88%, s.m. 5.9%, 
l.m. and tr. 4.7% and eos. 0.35%. There were also 4 normoblasts and 4 megaloblasts 
per 1000 leucocytes). 

In chronic tuberculosis a leucocytosis is the rule, especially if a secondary 
infection is present as usually is the case. Von Limbeck considers the 
presence of a leucocytosis proof that there is a secondary infection. Others 
disagree since in the chronic septicemia form, which is a pure infection, 
there usually is a slight leucocytosis and in caseous pneumonia (a pure 
infection) the leucocytosis may be as high as in croupous pneumonia. 

The normal count of the second stage has aroused considerable specu- 
lation. Some believe that potentially there must be an anemia but that 
this may be masked in some cases by the concentration of the blood due 
to sweating, diarrhea, vomiting or by a dyspnea which always tends to 
raise the count. Others think that there is an anemia which is covered by 
an oligemia and autopsies on patients with this stage of tuberculosis do 
suggest this. Von Limbeck claims that there is an oligemia due to a changed 
water metabolism which results in a general drying of the tissues and that 
this concentrates the blood. Grawitz says it is due to the lymphagogue 
effect of the products of caseous nodules. 

After hemoptysis the regeneration of the blood may be rapid (see page 
595). If after an operation on a tuberculous focus the hemoglobin does not 
rise rapidly the operation was probably incomplete. In some cases an ane- 
mia indicates an advance of the disease while in others the count may rise 
without any corresponding change in the condition of the patient. In 
fibroid phthisis there is as a rule no leucocytosis while in acute phthisis the 
anemia may be pronounced and progressive. In cases of cavity formation 
there usually is a leucocytosis. Some cases of extensive tuberculous pneu- 
monia have little leucocytosis; others, as high an one as croupous pneumonia. 
In acute miliary tuberculosis there usually is no change in the red blood- 
cells, hemoglobin or leucocytes but a few cases were reported with a very 
low white-count, even from 500 to 600 cells, over 90% of which are poly- 
morphonuclear neutrophiles. 

Tuberculosis of the serous membranes is accompanied by a mild secondary 
anemia (unless the blood be concentrated by diarrhea) without a leuco- 
cytosis except in meningitis, in which a leucocytosis is the rule (Osier). In 



THE BLOOD 637 

tuberculosis of the lymph-glands there is no leucocytosis, more often a leu- 
copenia, until caseation begins. 

The injection of tuberculin into a tuberculous patient sometimes causes 
a leucocytosis with a rise of eosinophiles. In tuberculosis of the bones 
there is marked absence of leucocytosis until a secondary infection de- 
velops ; a high white-count indicates acute abscess formation, but after the 
abscess has persisted for some time the count may remain normal until a 
secondary infection develops. In bone cases the red counts are rarely 
diminished, but the hemoglobin is low. 

That the anemia found in children should be so slight is rather remark- 
able since their blood usually is so susceptible to infection. Brown 142 
in 73 cases in very young persons found the red blood-cells diminished only 
in the long-standing extensile cases, but the hemoglobin was diminished 
somewhat in all. 

Of 17 cases of acute miliary tuberculosis in 5 the red cells stood between 3,600,000 
and 4,000,000 and in 6 over 5,000,000. The color-index was quite low, in y 2 the cases 
from 0.4 to 0.6. The leucocytes varied from 1000 to 9000, the majority (9) from 3000 
to 6000. One case had an interesting differential count (total, 3500; s.m. 6.5%, l.m. 
io.8%,pmn.n. 81.9%, eos. 0.5%). Warthin's case withalower count, had 91.48% pmn.n. 

Warthin reported a case with the leucocytes below 2,000, on 1 day 
(with a chill) 600, and Cabot a case with 550 white cells per 1 c.mm. 

In miliary tuberculosis, if in any condition, one would expect to find 
Bacillus tuberculosis in the blood stream (see page 29). How difficult this 
is, is shown by Wilson 143 in Warthin's laboratory who calculated from a 
study of the post-mortem clots from the heart that the heart blood at 
death contained no more than 40 individual bacilli. Dieter le, (ibid.) in that 
same laboratory, studying a case of acute miliary tuberculosis complica- 
ting chronic leukemia found one bacillus to each 15 slides of blood from 
the right heart. 

Secondary infections of the blood stream, however, are common in 
tuberculosis and increase in frequency as the disease advances. 144 Far-ad- 
vanced cases are 2^ times as likely to have them (61%) as are moderately 
advanced cases (24%) and these latter are 2^ times as likely as are incipient 
cases (9%). Febrile cases are 6 or 7 times as likely to have secondary 
organisms in the blood as afebrile cases, and open cases (40%) are 8 times 
as apt as closed cases (5%) to show organisms in the blood. 

Tuberculous Meningitis presents an illustration of the general rule that 
the tendency of an infection to cause a leucocytosis depends on the location 
of the lesion as well as on the organism, for tuberculosis of the meninges 
usually causes a leucocytosis. 

142 Trans. Med. Soc. of the State of California, 1897, p. '168. 

143 Jour, of Infect. Dis., Aug., 1916, Vol. 19, p. 260. 

144 Brown, Heise and Petroff, Tr. of the Ninth Annual Meeting of the Assoc, for 
the Study and Prevention of Tuberculosis. 



638 CLINICAL DIAGNOSIS 

In only 3 of 15 cases was the count below 10,000. One of these was 
a case of acute general miliary infection and in the other 2 but 1 count was 
made. The highest was 26,800. The leucocyte curve is a very irregular 
one, 3 of our cases with the high counts had also periods with low counts. 

In the series of 43 cases reported by Cabot there was a leucocytosis in 32. 

Tuberculous Peritonitis. — Of 19 cases of tuberculous peritonitis, 7 had 
counts between 3,000,000 and 4,000,000 and 6 between 4,000,000 and 
5,000 000. The color-index varied from 0.5 to 1. Of 22 cases there was a 
leucocytosis in 9 (highest count, 22,400). Of Cabot's 60 cases there 
was a leucocytosis in 14. 

Tuberculosis of Bones and Joints. — Of 15 cases of joint and bone tuber- 
culosis in the Johns Hopkins surgical clinic, 6 had a leucocytosis. 

It is believed that during the process of abscess formation a leucocytosis 
is the rule and that in time this will disappear to reappear in greater degree 
if a secondary infection develops. This may explain the high count follow- 
ing operation. This leucocytosis soon subsides and so long as the abscess 
drains freely will not reappear. 

Tuberculosis of the Intestine. — Of 5 cases of intestinal tuberculosis there 
was a secondary anemia in 2 (3,000,000 and 2,800,000 red cells) and in one 
a leucocytosis of 14,000 which disappeared soon after admission. 

Two cases of Renal Tuberculosis showed no leucocytosis. 

In one case of Addison's Disease the red cell-count was 6,000,000, 
hemoglobin 92% and leucocytes 9000. The rule in this disease is a 
marked anemia. 

Typhoid Fever. — For the bacteriology and serology of typhoid fever see 
pages 562 and 564. In the fresh blood smear in typhoid fever may some- 
times be found the very large phagocytic cells crowded with red corpuscles 
which Mallory described. Thayer 145 who reported the cases of the Johns 
Hopkins clinic found that from the end of the first week until defervescence 
there was a gradual reduction in the number of red cells and that regeneration 
began with defervescence. In very long-continued cases the regeneration 
may begin slightly before the temperature reaches normal. The total loss 
of red cells averages 1,000,000 cells. The average count at the end of the 
third week, which is the usual limit of the disease, was 4,555,814. The loss 
may be accentuated during the fourth week and, indeed, the usual statement 
is that the anemia begins at this time. Transitory variations are common 
during the fever due to vomiting, sweating, diarrhea, etc. After a severe 
hemorrhage the anemia may be marked and regeneration begin at once. 

Following some very severe cases a severe post-typhoid anemia may de- 
velop. In one case the red count was 1,426,000 during the fourth week; 
in another 1,300,000 during the third week (both Osier's cases) and in one 
804,000 (Henry). 

145 Johns Hopkins Hosp. Rep., vol. viii. 



THE BLOOD 639 

In one case, with distinct invasion of the bone-marrow by Bacillus typhosis, the red 
cell-count in 7 days fell from 3,752,000 to 1,006,000, the leucocytes rose from 4,200 to 
33,300 while 26,000 nucleated reds per c.m.m. appeared of which 2,880 were megaloblasts 
and intermediates. 

Usually there are no qualitative changes. After a hemorrhage nucleated 
reds are sometimes seen. 

There is always a more marked reduction in the hemoglobin than in the 
red cell-count and it returns to normal more slowly than do they. 

Some think that the leucocytes are slightly increased at the very onset 
of an attack of typhoid fever but most observers find them subnormal dur- 
ing the whole course. Certainly their count diminishes gradually from the 
end of the first (when the average count is 5400) to the fifth week, at which 
time the average of Thayer's cases was 5386. Some cases reach 2,000, 
others 1,000 per c.mm., some even lower. Thayer found no cases with an 
initial leucocytosis. The longer and more intense the infection the lower 
the leucocyte count. In a few cases without complication the count is 
above 10,000 throughout the whole course. Temporary variations are 
common, the count rising, e.g., to 10,000 cells after a cold bath, yet 
with the differential count unchanged. 

The differential count for the first 5 weeks shows a progressive decrease 
in the polymorphonuclear neutrophiles, usually to 60%, often below 50%. 
There is also an increase of the endothelial cells. These are especially nu- 
merous at the height of the fever. The eosinophile cells are below 1% as a 
rule until convalescence, when they increase even to an eosinophilia. They 
may, however, in long continued cases, rise with the increase in the 
red cells before the temperature is normal. During convalescence the 
count slowly rises, but the blood retains its characteristic features for 
about 3 weeks after the temperature is normal. 

The blood picture maybe modified by various complications . Hemor- 
rhage causes an acute post-hemorrhagic anemia with leucocytosis. The low- 
est count of Thayer's series was 1,992,000 cells. Regeneration begins at once. 

The inflammatory complications cause a rise of the count of white cells, 
even a true leucocytosis. This is true of furunculosis, phlebitis, thrombosis, 
bronchitis, periostitis, pleurisy, pneumonia, etc. A definite rise of a count 
already very much reduced is for that person often a true leucocytosis ; for 
instance, one case of typhoid fever with a leucocyte count of 1600 developed 
parotitis whereupon the count rose to 3200 cells, a reaction comparable in a 
normal person to a rise to about 15,000 cells. In 1 case with empyema the 
count was 44,500; in a second case of empyema due to Bacillus typhosus the 
count was 23,000 cells of which 68.5% were polymorphonuclear neutro- 
philes, 12.7% small mononuclears and 17% large mononuclears. 

Three of 5 cases of pneumonia, of whom 2 died, had counts above 10,000 
cells and 2 cases, both of whom died, had counts below 10,000 cells. The 
counts in all of these cases had a rather small percentage of polymorphonu- 



640 CLINICAL DIAGNOSIS 

clear neutrophiles. In i case of periostitis due to Bacillus typhosus the 
leucocytes were 18,000 of which 72.5 % were polymorphonuclear neutro- 
philes. Thayer cites many similar illustrations showing that in typhoid 
fever the leucocyte reaction in cases with complications depends much 
upon the tissue infected and that there is a tendency, in cases with a 
leucocytosis for the formula of typhoid fever to persist. 

In all cases of suspected intestinal perforation the leucocytes should, 
from the first, be followed with the greatest care. In most cases the leu- 
cocytes rise either to an absolute leucocytosis of 10,000 or over, or to one 
relative to the previous counts. Then the count sometimes drops coincident, 
perhaps, with the spread of the peritonitis. In some fulminant cases the 
count falls from the first. In the so-called pre-perforative stage there is a 
slight leucocytosis due to the local peritonitis. While the leucocyte curve 
has but little absolute value, yet it has much when interpreted in the light 
of the physical examination. If the abdominal signs are suggestive of 
perforation the operation is performed whatever the leucocytes may 
show. But if the local abdominal signs alone seem hardly sufficient 
to justify operation a rising count would settle the question. Of course a 
rising leucocytosis might mean something other than perforation. In one 
of our cases it meant appendicitis. Every case in which there is very good 
reason to fear perforation is operated upon, under the belief that a quite 
unnecessary operation will be of benefit since if perforation has occurred 
from 30 to 50% will be saved and if there is no perforation the course of the 
fever will at least probably be milder after the operation. 

Pneumonia. — In acute lobar pneumonia the coagulation of the blood is, 
as a rule, rapid. The count of the red blood-cells is normal during the fever, 
but there may at first be a rise, as in Sadtler's case to 7,000,000. After the 
crisis there is always a drop of about 500,000 cells and sometimes a slight 
post-febrile anemia. The hy^ercythemia is probably due to a concentration 
of the blood which may cover a real anemia caused by the loss of blood in 
the exudate and by the destruction of the blood-cells, as shown by the 
jaundice and the urobilinuria. Some cases have a true and severe anemia 
with a loss of about 2,000,000 cells. On the day of crisis there is a drop due 
in part to a general peripheral relaxation (Grawitz) and partly to a true 
anemia which up to this time has been covered by the hypercythemia. 

Nucleated reds are more common in the blood in pneumonia than in 
other acute fevers. Both normoblasts and megaloblasts may be present. 
The latter have a bad prognostic import only when present in considerable 
numbers. It is thought that at the time of the crisis the cells crenate 
more readily than normal. 

In 34 cases of acute lobar pneumonia studied with special reference to the red 
blood-cells there was a drop in the count during the lysis or just after the crisis, generally 
of about 1,000,000 but in some cases of 2,000,000 cells, which usually only restored the 



THE BLOOD 641 

count to that level which obtained before the hypercythemia. The later counts showed 
small gains and losses in an even number of cases and of about the same degree, but in 
9 cases there was a permanent loss of from 900,000 to 1,500,000 and in 4 cases a gain of 
from 700,000 to 1,900,000 cells. 

In pneumonia the inflammatory leucocytosis has been best studied. 
None of our cases showed an initial hypoleucocytosis, as claimed by Pick. 
From the first, i. <?., from 6 or 8 hours after the chill, the leucocytes are in- 
creased. The maximum count just precedes defervescence and the drop 
immediately follows this. This leucocytosis is an expression of the resist- 
ance of the patient to the infection and depends but little on the fever or on 
the extent of consolidation. Cabot has divided the cases into 3 groups : (1) 
Those with good resistance and a mild infection, in which there need be no 
leucocytosis; these cases all recover. (2) Those with a severe infection and 
a good resistance, in which the leucocytosis is high, between 20,000 and 
30,000, but in some cases over 100,000 and even 1 1 5,000 (Lohr) . This group 
includes about 90% of all cases. (3) Those cases in which the infection is 
severe and the resistance poor. In these there is no leucocytosis or even a 
fall. These cases are usually always fatal. This last group includes the 
terminal pneumonias of chronic diseases, the pneumonia in the aged, etc. 
In fatal cases the percentage of polymorphonuclears may rise considerably 
although the total count may not at all. 

The statement is made that the leucocytes do not drop with a pseudo- 
crisis and even rise. The fall in the leucocytes begins just before, just after 
or with that of the temperature. In cases ending by crisis the count falls 
by lysis reaching normal on about the second day, while if the temperature 
falls by lysis the leucocytes fall still more slowly. If a slight temperature 
persists after the crisis the leucocytes remain elevated until the temperature 
is normal. In fatal cases there is often an ante-mortem rise of the white 
cell count. If there is delayed resolution the leucocytes may stay elevated 
even for weeks and then slowly drop with the temperature. For the count 
to remain elevated after a supposed lysis suggests delayed resolution, 
empyema, or pulmonary gangrene. 

A high leucocyte count has no value in prognosis ; it merely means that 
the patient is making a vigorous fight. 

In the Johns Hopkins Hospital the leucocytes of pneumonia cases were counted 
twice daily. We 146 have compiled the records of 158 uncomplicated cases with recovery, 
56 uncomplicated cases with death and of 80 cases with various complications. In the 
uncomplicated cases with recovery the degree of the leucocytosis bore no relation to the 
extent of the consolidation. In 38% of the uncomplicated cases with recovery the 
count was below 20,000 and in 7% above 40,000. Age had little influence on the leuco- 
cyte reaction since exactly the same percentage of cases below 40 years of age had a 
leucocyte count below 20,000 as of those older. 

Of the 77 cases who terminated by crises the leucocyte counts of those above 40 years 

146 Johns Hopkins Hospital Reports, 19 10, Vol. XV. 
41 



642 CLINICAL DIAGNOSIS 

of age were somewhat higher than of those younger. In 18% of these cases the counts 
varied from 10,000 to 15,000. These, clinically, were very mild cases. In 25% they 
varied from 15,000 to 20,000, and in 8% were over 40,000. There was a sharp precritical 
rise in 42% of these cases. 

Of 81 cases with lysis the count during the course kept below 10,000 in .2%, from 
10,000 to 15,000 in 20%, from 15,000 to 20,000 in 14%, and above 40,000 in 10%. There 
was a sharp rise just at lysis in 34% of these cases. 

Those rises which occurred just before lysis or crisis amounted to from 5000 to 10,000 
cells as a rule, but in a few it was over 20,000, and in 1 case 30,000. The highest count 
seen in a case was 105,500 in a young man 25 years old who recovered. 

Of the cases with crisis the fall in the leucocytes preceded the drop of temperature 
in 15%, accompanied it in 41% and followed it in 44%. In the cases ending by lysis 
the drop began before that of the temperature in 18%, with it in 43%, and followed 
it in 39%. (Notice the similarity in these figures.) For the leucocyte count to reach 
normal required, in the cases with crisis, from 1 to 20 days; mean, 3 days. 

A well-marked pseudocrisis occurred in 9 cases. Of these 2 were accompanied by a 
rise of leucocytes, 4 by a fall, and 3 by no change in the white cell count. In cases in 
which a slight fever continued after its drop the leucocyte count remained from 12,000 
to 15,000 until the temperature reached normal. 

There were, in this series, 56 fatal cases. The leucocyte counts in these were almost 
the same for the various decades of age as in those with recovery. During the course 
they remained below 10,000 cells in 23 of the cases (in 1 case they reached even 1700); 
from 10,000 to 15,000 in 23%, from 15,000 to 20,000 in 15% and over 40,000 in 1 case. 
At the time of death the count was below 10,000 in 17%, from 10,000 to 15,000 in 25%, 
15,000 to 20,000 in 10% and above 40,000 in 8%. All of the last cases were under 30 
years of age. Toward death there was in 70% a progressive rise and in 30% a fall. 

The absence of a leucocytosis does not necessarily indicate a fatal outcome. In 1 
case with extreme toxemia and a count of 8000 the leucocytes slowly rose to 14,000 as 
the patient recovered. 

Daily Variations in the Count. — Counts made in the forenoon and afternoon, separ- 
ated by an interval of about 9 hours, differed by from 1000 to 26,000 cells (as a rule from 
4000 to 6000; mean, 4000). There was no difference in these variations before and after 
the crisis. While the temperature is fairly constant the counts vary less and yet there 
was in general no parallelism between fluctuations of temperature and of leucocytes. 

In cases of delayed resolution the leucocytes reached normal before, with, or after 
the temperature. In some cases both temperature and leucocytes were normal before 
resolution was complete. 

The cases of terminal pneumonia presented great variations. In our series there 
were 2 counts above 50,000 and 2 below 3500. Alcoholics had almost no leucocytosis 
and yet some recovered. In no cases followed by empyema did the leucocyte counts 
indicate the onset of this sequela. In 1 case of pneumonia followed by empyema the 
leucocytes did not rise at all until the empyema began while in another such case they 
had not risen up to the day of the operation. In 2 cases followed by pleurisy with 
effusion the leucocytes were normal after the crisis (6000 and 8000). In 3 fatal cases 
ending in abscess of the lung the leucocytes were respectively 46,000, 30,000 and 8500. 
In 35 cases with various pus infections, endocarditis, pericarditis, meningitis, parotitis, 
otitis media, phlebitis, thrombosis, tonsillitis, etc., very little could be learned from the 
leucocyte counts, i.e., they did not change with the development of the complication. 

Qualitative Changes. — The leucocytosis of pneumonia is of the polymor- 
phonuclear neutrophile variety and yet the percentage of these cells is 
seldom 90%, often it is not over 80%. After the crisis it may drop to 60% 



THE BLOOD 643 

and even lower, due in part to an absolute increase of mononuclear non- 
granular cells. The eosinophile cells may disappear from the peripheral 
circulation during the attack and reappear at the crisis, at which time also 
myelocytes, even 12% of the total white cell-count, may appear. The large 
basophile mononuclears also may be increased. For the percentage of 
polymorphonuclear neutrophile cells to be above 90 or below 50 is thought to 
indicate a bad prognosis. 

Glycogen can usually be demonstrated in the leucocytes, in amounts 
varying with the temperature and the extent of consolidation. 

The platelets may even disappear during the fastigium but after the 
crisis they return and increase to above normal. 

The fibrin network is much increased. Coagulation is rapid. The 
specific gravity of the blood varies as the count and is high. The toxicity 
of the blood is even doubled. 

In a doubtful case a high leucocytosis would exclude malaria and typhoid 
fever and suggest a central pneumonia. This is especially important in 
the very old and in the very young patients. 

In the bronchopneumonia of influenza of the epidemics of past years 
there has been a polymorphonuclear neutrophile leucocytosis of from 
10,000 to 15,000, and in severe cases of from 20,000 to 25,000 cells per c.mm. 
147 but during the epidemics of 1917-20 it was more common to find a 
leucopenia of under 5000, i. e., 2800 or below, even in cases which 
develop an empyema. In one of our cases it was 83,400. 

In acute epidemic cerebrospinal meningitis there is a leucocytosis (see 
page 513) but this disease is not primarily an inflammation of the meninges 
but is a septicemia in which the bacteria tend to become localized in the 
membranes of the brain and cord. This is what might be expected from the 
clinical picture, especially that of the fulminating cases. That the bacteria 
must frequently be present in the blood in large numbers and at a very 
early stage is suggested by the very early appearance of the petechial 
eruption (hence the term "spotted fever"). 

In cases of infection by intestinal parasites a slight leucocytosis is the 
rule. In 12 of our 18 recent cases these cells varied from 11,200 to 34,000. 
In 4 cases with fever the counts were normal. 

Bronchial Asthma. — In bronchial asthma the most interesting find is an 
eosinophilia of even 53.6%. This is important in diagnosis and also as a 
means of predicting oncoming paroxysms (see page 522). 

Of 17 cases, the red cell count was over 5,500,000 in 7 and the lowest count was 
4,900,000. There was a leucocytosis of from 10,000 to 15,700 in 6 cases. Of 8 cases in 
which differential counts were made 6 had an eosinophilia. (The absolute numbers 
of eosinophiles in these cases were 728, 712, 535, 856, 702 and 1720 [20% of 
8600 leucocytes per 1 cmm.]). 



Davis, Arch. Int. Med., 1908, vol. ii, p. 124. 



644 CLINICAL DIAGNOSIS 

Acute Rheumatic Fever. — " The blood is the best index of the severity 
of this disease" (acute rheumatic fever) (Osier). Its virus causes rapid 
destruction of the red cells, lowering the count often (but not always) 
from 1,000,000 to 2,000,000 cells. The high counts sometimes seen during 
the attack may be due to the profuse sweats. The anemia is most evident 
at the time of convalescence. Hay em, Turk and others say that the count 
is lowest at the height of the fever and that regeneration begins at once 
with defervescence. Nucleated reds are seldom present, In no other 
disease is the fibrin network so thick. 

There is a leucocytosis as a rule, the count running parallel to the se- 
verity and acuteness of the disease. Cabot's average was 16,000. The 
blood formula is that of an acute inflammatory disease. 

Of 77 cases of the Johns Hopkins clinic the red cell-counts varied from 2,000,000 to 
3,000,000 in 3, from 3,000,000 to 4,000,000 in 15, from 4,000,000 to 5,000,000 in 45 and 
5,000,000 and over in 14 cases. The mean count was 4,500,000. In these cases therefore 
there was very little anemia. Of 81 cases, the leucocytes were below 5000 in 1 case, from 
5000 to 10,000 in 23, 10,000 to 15,000 in 36, 15,000 to 20,000 in 15 and above 20,000 
in 6 cases. 

One case, a man 56 years of age, was admitted with red cells 1,720,000, hemoglobin 
27% and leucocytes 12,400. He gave the history of painful, swollen, red joints 4 weeks 
before. He recovered rapidly. 

The bacteriological study- of the blood has failed to throw any new light 
on the etiology of this disease and the opinion is still held by many that if 
any organism can be cultivated from blood or joint the case is one of 
acute infectious arthritis and not of acute rheumatic fever (acute articu- 
lar rheumatism) . One of the last good studies of this condition is that 
of Swift and Kinsella, 148 who found no type of streptococcus constantly 
or frequently (i. e., not over 10%) associated with this disease and those 
which were found varied so much that none deserved the name Strepto- 
coccus rheumaticus. 

In subacute or chronic rheumatism there is no leucocytosis. 

Arthritis Deformans. Subacute Infectious Arthritis. — McCrae found 
that in 33 cases of arthritis deformans the average of hemoglobin was 
70.6%, of the red cells (in 29 cases) 4,468,000 and of the leucocytes, 7600. 
The differential counts were normal. 

Appendicitis. — In cases suggesting acute appendicitis the leucocytes are 
counted each hour. If with suggestive history and symptoms there is a 
rising leucocytosis an operation is performed without delay, while if the 
abdominal signs are marked the operation is performed whatever the leu- 
cocyte count may be. If the leucocyte counts are stationary, even though 
high, when the patient is first seen, one may wait ; but if rising even slightly 
there should be no delay. The count may be normal in mild or very severe 
cases, or in cases with well walled abscess. A leucocytosis of 20,000 or above 

148 Arch, of Int. Med., Mch., 1917, xix, p. 381. 



THE BLOOD 645 

indicates acute appendicitis, probably an appendix full of pus and quite 
tense. This will fall after the appendix ruptures. (At least those cases ad- 
mitted soon after the appendix had ruptured often have low or even sub- 
normal counts even though the peritonitis is spreading.) In appendicitis 
a count above 15,000 means an active process. Over 20,000 is a high 
count and means pus, gangrene, or peritonitis. Fulminating cases may die 
without any sign of reaction on the part of the white cells. 

In chronic appendicitis with abscess a stationary leucocytosis means 
that the latter is well walled off. If the abscess has been present for some 
time the count is seldom above 12,000 and usually is nearer 7000. If now 
one operates the count will at once rise to 20,000 or over and then gradually 
drop. This is perhaps due to the exposure of new tissue to infection or to 
absorption. If the count after the operation remains high it means that a 
pus pocket is still unopened. In cases with well- walled abscess and a normal 
leucocyte count the leucocytes may fluctuate markedly. For this no ex- 
planation can yet be offered. In cases with a spreading peritonitis the count 
of white cells may rise, or drop and then rise, or may fall even to subnormal. 
The falling leucocytosis is a worse sign than a high stationary count. 149 

In CHRONIC OBLITERATIVE APPENDICITIS and SUBACUTE APPENDICITIS 

without exudate there is no leucocytosis. 

The red cells are not affected in cases of appendicitis unless there is a 
long-standing abscess, then there may be a secondary anemia. Da Costa 
mentions an early slight anemia in most cases, in some a severe anemia. 

ANEMIAS OF CHILDREN 

The study of the blood of normal children is very important since they 
react to disease often differently from adults. We have even known a 
diagnosis of lymphatic leukemia suggested by a normal child's blood. 

Children are much more susceptible to the agencies which produce 
anemia than are adults, their anemias develop more rapidly, become more 
severe and have a worse prognosis. 

In the anemias of the very young a lymphocytosis is usually present 
and unripe elements soon appear, striking evidence of the activity of the 
bone -marrow. Among these are normoblasts, megaloblasts, myelocytes and 
large basophilic non-granular leucocytes which do not appear in normal 
blood. But these qualitative changes in the blood picture have less sig- 
nificance than in the adult and merely show the greater instability of the 
blood-regulating mechanism. 

Certain cases of anemia are grouped under the term "anemia of 
growth" and are said to be due to the inability of the blood-building organs 
to keep pace with the increasing demands of the growing body. During 
this period wasting diseases, infections and any agencies deleterious to the 

149 See Bloodgood, Prog. Med.. December, 1901. 



646 CLINICAL DIAGNOSIS 

blood, vascular system, or to the hematopoietic organs, such as poor food 
and bad hygienic surroundings, have a more pronounced effect than on the 
blood of an adult. This is well illustrated by the anemias of school children, 
which are ascribed by some to mental strain, lack of exercise, poor appetite, 
constipation, etc., but by others to bad tonsils, latent tuberculosis, etc. 
The development of the heart and blood vessels is closely related to that of 
the blood, hence chlorosis and other severe anemias of youth occur in as- 
sociation with hypoplasia of the cardiovascular system ancl are attributed 
to congenital defects. 

A severe anemia in a child is perhaps never perfectly recovered from. 
Objective evidence of it may disappear and the child seem well, but rela- 
tively insignificant illnesses will bring it to light again. 

The Anemia Pseudoleukemica Infantum of v. Jaksch was described 
as a severe anemia of young children, with the red cell-count usually from 
1,500,000 to 3,500,000 but even as low as 820,000, a low color-index (0.50) 
and a leucocytosisof even 54,660 (in one case, 1 14,000) withafewmyelocytes 
present. Many of the red cells are deformed and degenerated and many are 
nucleated. The leucocytes are characterized by their great variations in 
form, size and staining qualities. The platelets are increased in number. 
Cabot 150 thought that many different diseases are grouped under this 
diagnosis, including pernicious anemia, secondary anemia with leucocytosis 
(due especially to lues, rickets, etc.), Hodgkin's disease and even leukemia, 
all of which' diseases are apt to be atypical in children. 

Malaria of children may cause a severe anemia with normoblasts and 
practically always megaloblasts present, but no marked leucocytosis unless 
it be due to an increase of the endothelial leucocytes. 

Congenital lues causes the severest anemias. Many nucleated reds 
appear, the lymphocytes especially are much increased and the total white 
cell-count may reach even to from 50,000 to 100,000. 

Rickets causes a simple chlorotic anemia with a leucocytosis of even 
30,000 which is in large degree a lymphocytosis. 

A case of anemia in a 14-months-old child is entered on our records sim- 
ply as "anemia with enlarged liver and spleen " (Osier). The red cells on 
admission numbered 1,252,000 per c.mm., the hemoglobin 20% and the 
leucocytes 14,700. The child was in the ward 'one month without improve- 
ment. The leucocytes varied from 13,000 to 26,500, always with the same 
formula (s.m. 40 to 52%, l.m. and tr. 5 to 18%, pmn. n., 38 to 62%, eos. 
o to 0.9%, neutroph. myeloc. 0.9 to 3% and Mastzellen o to 0.2%. The 
nucleated reds, which numbered from 24 to 250 per 1000 leucocytes, were 
chiefly normoblasts, some were intermediates and some megaloblasts and 
microblasts). Such a case resembles the French " splenomegalie chronique 
avec anemie et myelemie." 

150 " Clinical Examination of the Blood," Fifth edition, p. 519. 



THE BLOOD 647 

Slimmer Diarrheas of Children. 151 — In some cases of severe diarrhea in 
children the red cell-count may rise even to 10,000,000 cells. The ordinary 
summer diarrheas are usually accompanied by a leucocytosis. In the 
simple dyspepsias the differential count of leucocytes is normal (total 13,500 
to 36,000; s.m. 39%, l.m. and tr. 21.2%, pmn. n. 37.8% and eos. 2%), but 
in the more severe cases there is an increase in the polymorphonuclear 
neutrophiles (from 56 to 63%) and a decrease of the mononuclear cells 
(from 33 to 7%), the blood thus presenting the adult formula. In an acute 
intestinal toxemia and in the severe forms of enterocolitis a true leucocy- 
tosis is the rule, which is characterized by the presence of a few myelocytes 
and the absence of eosinophiles. 

A leucocytosis with a wasting disease in a child usually indicates an 
inflammatory intestinal complication. 

CHRONIC DISEASES 

Chronic Nervous Diseases. — The blood in cases of chronic diseases of 
the nervous system presents nothing at all characteristic. Whatever 
changes are present depend on the general nutritional condition of 
the patient. 

Such patients with Acute Chorea are very anemic, yet of 23 cases of 
the Baltimore Clinic this was true of but 3 and these 3 had heart compli- 
cations. The lowest count was 3 ,400,000 reds. These cases sometimes have 
an eosinophilia of from 7 to 10% (Theleme). 

In general paresis 152 Capp and Jenks found in some cases just before 
a paretic seizure an absolute leucocytosis, the increase affecting especially 
the large mononuclears. A slight anemia which progresses with the dis- 
ease is the rule, except during the seizures when the count of red cells may 
temporarily rise. 

In Maniacal Depressive Insanity 153 an anemia is the rule and al- 
most always during the periods of excitement there is a leucocytosis. 

In Acromegaly there is usually an increase in the count of red cells 
with eosinophilia and lymphocytosis (Ducati). 

Diabetes Mellitus. — In diabetes mellitus one of the essential symp- 
toms during the periods of glycosuria is the hyperglycemia of even 
0.57% instead of, as normal, 0.1 to 0.2%. This hyperglycemia may 
cause a hydremia, i. e., sl dilution of the blood, and therefore a lowering 
of the count of red cells while the resulting diuresis tends to concentrate 
the blood. Later in the disease a cachexia with anemia develops which, 
however, may be well masked by the concentration of the blood. The 
leucocyte counts are normal. The patients often show a remarkable diges- 
tive leucocytosis. 

151 Knox and Warfield, John Hopkins Hospital Bull., July, 1902. 

152 Am. Jour, of Insanity, January, 1900; Diefendorf, loc. cit., 1903, vol. cxxvi. 

153 Fisher, Am. Jour. Insan., April, 1903. 



648 CLINICAL DIAGNOSIS 

Of 45 cases, the red cell-counts were below 4,000,000 in 3 cases (the lowest count 
was 2,000,000), from 4,000,000 to 5,000,000 in 13, from 5,000,000 to 6,000,000 in 10 and 
over 6,000,000 in 4. In 3 other cases the count at times was over 6,000,000. 

Of 40 cases, the leucocytes varied from 5,000 to 10,000 in 25, from 10,000 to 20.000 
in 7 and over 20,000 in 7. The highest leucocyte count was 44,000. These leucocytoses 
were due to pneumonia, septicemia, furunculosis, gangrene, etc. In a case of Coma the 
leucocytes varied from 30,000 to 41,000. 

Blood Lipoids. — "With an excess of fat diabetes begins and from an ex- 
cess of fat diabetics die" is the very expressive way Joslin emphasizes the 
study of blood lipoids in diabetes. Lipemia (see page 556), or the pres- 
ence in the blood of sufficient visible fat to give the plasma a milky appear- 
ance, is common in severe cases of diabetes on a fat-rich, carbohydrate- 
poor diet, but usually disappears after the diet is changed. 

Basal Metabolism. — The conclusions of the accurate work of Joslin, 
Benedict and their co-workers on basal metabolism in diabetes may be 
summarized as follows : 

While even in the same patient metabolism may at one time be increased 
and at another be below normal, yet sevei e cases of diabetes during the 
existence of an acidosis show an increase in metabolism of from 15 to 
20%, the increased consumption of oxygen and the increased heat elimi- 
nation running hand in hand. A similar increase is seen in normal per- 
sons in whom an acidosis is artificially induced. Following the disappear- 
ance of the acidosis the metabolism may become normal or below normal so 
that severe diabetics may live on less calories than the normal individual. 

Bremer 1 s blood-test for diabetes mellitus has, in some cases, proved of value. A 
thick smear of blood on a slide and a similar one of normal blood for a control are sub- 
jected to exactly the same treatment They are first heated to 135 C, then allowed to 
cool slowly and are then stained for 2 minutes with 1 % aqueous Congo red solution. The 
diabetic blood will take a yellower stain than the normal. This test is said to be positive 
when the urine is sugar-free and even before sugar has ever appeared. Schneider found 
it positive in the cases of 2 normal men who were great meat-eaters and ascribed it 
to the reaction of the blood. Strauss confirms this opinion, finding it best in cases 
of acidosis. It is claimed to be sometimes present in leukemia, Hodgkin's disease and 
Graves' disease. 

In this clinic a man was admitted during coma; no urine could be obtained by 
catheterization; the diagnosis of diabetic coma was made from this test alone and was 
confirmed later at autopsy. 

Williamson's Test. — Twenty cubic centimeters of blood in a test tube are mixed 
with 1 c.c. of aqueous methylene blue (1 to 6000) ; 40 c.mm. of 6o%KOHand40 c.mm. 
of water are added. This mixture is allowed to stand for 3 or 4 minutes in boiling water. 
If the blood is diabetic it takes a yellow color. 

Malignant Disease. — Malignant tumors are among the most important 
of anemia-producing diseases. They certainly produce a toxin which injures 
the blood, but the hemorrhages and the mechanical effects of the cancer 
on the gastro-intestinal functions must not be overlooked . And yet it is 



THE BLOOD 649 

remarkable how long the blood will remain almost normal and then how 
rapidly cachexia and anemia will develop. 

Cancers differ much in their effect on the blood. Some, for instance 
those of the skin or lip, may cause none or a slight secondary anemia, the 
so-called "pseudo-chlorosis carcinomatosa, " while one of the stomach, 
''may give the perfect picture of primary pernicious anemia or, indeed, 
of leukemia." It is stated that the more malignant the tumor and the 
more extensive its metastases, the greater its influence upon the blood. 
But this certainly is not true. Our cases of huge tumors with rapidly 
spreading metastases had a slight chlorotic anemia while those which 
simulate pernicious anemia are more apt to be an insignificant-looking 
little nodule, usually on the stomach and discovered at autopsy. Those 
which cause the severest anemia are those which give rise to frequent 
hemorrhages, e. g., those of the stomach and uterus and those also 
which mechanically disturb the functions of the digestive tract. In many 
cases with advanced cachexia, yet no hemorrhages, there may be even 
a rise in the red cell-count due to desiccation of the tissues (v. Limbeck). 

The anemia due to cancer is as a rule of the secondary type and severer 
than that due to any other chronic disease. The first changes are in the 
size, shape and weight of the red blood-cells and later in the counts. As the 
cachexia develops the red cell-count may be as low as 2,500,000 cells or even 
1,000,000 cells. An exception to this is in cancer of the esophagus in which 
cases the blood may be concentrated. Grawitz suspects that in some cases 
the anemia is in part only apparent, since the injection of carcinoma extract 
will produce a hydremia. There is a constant and often an early re- 
duction in the hemoglobin. If it is normal the cell count will be found above 
normal. The average in long-standing cases is about 68.5%, in worse cases 
57-5%, while the color-index averages about 0.65. This low color-index is 
an important diagnostic point between malignant and non-malignant 
tumors. Later, if the anemia assumes the pernicious type, the color-index 
may rise above 1. It has been claimed that after the removal of a cancer 
by operation the regeneration of the blood begins late and is never 
quite complete. 

While the anemia is of the chlorotic type many or most of the red cells 
will be diminished in size while later, after it assumes the primary type, the 
cells will be large; yet the giant cells of pernicious anemia will be rarely 
seen except late, while microcytes will be numerous (Grawitz). The baso- 
phile granulation is very common. The deformities in size and shape and 
the degenerations of the erythrocytes may be absent or they may even be 
more marked than in tuberculosis, in which case this would be of diagnostic 
value. In any case, however, they will be a less prominent feature than in 
pernicious anemia. Nucleated reds are found even when the anemia is slight 
and in greater numbers than in the secondary anemies due to other causes. 
Their presence has a limited value in the differential diagnosis between 



650 CLINICAL DIAGNOSIS 

cancer and ulcer of the stomach. There will be normoblasts as a rule al- 
though in those cases which simulate pernicious anemia a few megaloblasts 
also may be present. In cases with metastases to the bone-marrow their 
number may be surprisingly large. 

There is a moderate leucocytosis in about 60% of all cases of malignant 
disease, an important point in the differentiation between benign and ma- 
lignant tumors. This may be the first sign of a cachexia. Some cases sug- 
gest even a leukemia. In malignant esophageal stricture the starvation 
sometimes causes a leucopenia with relative lymphocytosis. Cancers of the 
uterus and stomach, so commonly accompanied by hemorrhage, usually 
cause a leucocytosis ; and in malignant disease of the thyroid, pancreas and 
kidney the count sometimes is especially high. It is said that the faster 
the tumor grows the higher the leucocyte count, but there are many 
exceptions to this rule. Grawitz, who explains the leucocytosis as due to a 
lymphagogue action of the cancer extract which sweeps a great many 
leucocytes from the tissue spaces into the capillaries, considers that any 
leucocytosis is coincident with the softening of a tumor mass. After oper- 
ation the leucocytes will drop, while a subsequent rise may indicate a 
recurrence of the disease even before it can be found physically. 

The majority of leucocytoses due to malignant disease are of the poly- 
morphonuclear neutrophile variety but some are lymphocytoses of even 
43.7%. In other cases with a leucopenia of only 3,000 cells even 88.7% 
of these are polymorphonuclear neutrophiles. The eosinophiles are usually 
more in evidence than in other leucocytoses. Myelocytes are more numer- 
ous in the blood of cancer patients than in all other conditions except 
leukemia and pernicious anemia. It is said that a cancer may cause de- 
generative changes of the leucocytes before the quantitative changes begin. 

The specific gravity of the blood is low. The plasma is rich in sugar, 
as rich even as in diabetes. Its alkalinity may be considerably decreased. 
The coagulability is normal or lessened unless sloughing or inflammation 
is present, in which case it may be rapid. The fibrin network usually 
is normal. 

Cancers of the breast usually cause a slight leucocytosis (e. g., leucocyte 
count 11,000). Cancers of bone often give a blood picture with many nu- 
cleated reds, both normoblasts and megaloblasts, and a leucocytosis with a 
high percentage of non-granular mononuclear cells and some myelocytes, 
but not as many coarsely granular cells as one might expect. 

Cancer of the Stomach. — Cancer of the stomach usually causes a 
rather marked chlorotic anemia. Cabot found that of 129 cases, in 27 the 
red cell-count was above 5,000,000, in 26 below 3,000,000 and that the aver- 
age of all cases on the first examination was 4,018,000. Of the 134 cases of 
the Johns Hopkins clinic, including those reported by Osier and McCrae, 
in 33 it was above 5,000,000 and in 16 below 3,000,000. The mean was 
about 4,000,000. The color index was always considerably below 1 unless 



THE BLOOD 651 

the count was very low. Nucleated reds were rather rare. The count some- 
times drops progressively till death (in one case to 1,786,000). High 
counts may sometimes be attributed to excessive vomiting. The differ- 
ential diagnosis between cancer of the stomach and pernicious anemia is 
one of well-recognized difficulty and in many cases can be settled only at 
autopsy. The red cell-count may be as low as 500,000, but such cases 
are rare. In general it may be said that in cancer there is less anemia than 
the cachexia present would suggest, while in pernicious anemia the reverse 
is true. It is often said that a count below 1,500,000 is against cancer, but 
this rule often fails. In cancer one may expect to find fewer nucleated 
reds than in primary anemia and those present are more apt to be 
normoblasts, while a leucocytosis is more common. The highest of our 
series was 52,800. The leucocytes vary much in cancer of the stomach. 
A leucocytosis is present in over one-third of the cases and in those with 
normal counts a digestive leucocytosis is often absent (in 82% of 144 
cases). Counts below 4000 are not rare. 

It is said that the rapidity of growth of a gastric cancer influences the 
leucocyte count and yet our lowest counts included those with metastases 
in liver, pancreas, or peritoneum (1600, 5400, 5000, 5600), while in 15 
cases of general carcinomatosis the leucocytes were above 10,000 in but 7. 
In one case the count was 105,000 (t.° 103 F.) ; in another, 24,500 (t.° 99 ) 
and in a third, just before death, 61,400. These high counts were met with 
nearly always in cases with a slight fever. 

Cabot reports a leucocytosis of 105,600 in a case with perforation into 
the peritoneum followed by quickly fatal peritonitis. We suspected this 
condition in a case the count of which rose to 120,000, an almost pure 
leucocytosis, but were unable to get an autopsy. 

The percentage of large mononuclears is often high (1 to 10%) 
while in one case before death it was 33% of a total count of 6300 
leucocytes (Kurpjeveit). 

In Carcinoma of the Esophagus the blood is apt to be concentrated. 
This raises the percentage of dried substances, as in v.Noorden's cases, 
to 26.5 and 27.3%. And even in these cases also there may be an oligemia. 
If the cancer extends to the larynx, causing dyspnea, a high count may be 
due to cyanosis. 

Of 6 cases, the highest red cell-count was 5,960,000 and the lowest 4,184,000. In 
another case on first blood examination the red cell-count was 4,696,000, hemoglobin 
85% and 6000 leucocytes. A later examination in this case gave 6,476,000, 104% and 
19,000 respectively. Five of the cases showed a leucocytosis. The highest count 
was 30,250. 

In 15 cases of general carcinosis of the abdominal organs, cases in which one might 
assume a rapid and extensive growth, in but 2 was the red count below 4,000,000. In 
27 cases of cancer of the bile-ducts the lowest red cell-count was 3,700,000 and in 5 of 
these the leucocyte count was above 10,000. (In 1 case it was 44» I 5°; t0 I00 ° F -) 



652 CLINICAL DIAGNOSIS 

In 4 cases of cancer of the rectum the lowest red cell-count was 3,732,000. The rest 
were about normal. There was a leucocytosis in 2 cases (13,100 and 19,750 ; t° ioo° P.). 

Of 10 cases of cancer of the intestine 3 showed a marked anemia. In 1 , a cancer of the 
ileum, the red cell-count was 1,600,000, hemoglobin 40% and leucocytes 2500; in another 
the red cell-count was 1,780,000, hemoglobin 28% and leucocytes 10,000. This patient 
had nephritis also. In the third case, with cancer of the sigmoid flexure, the red cell-count 
was 1,609,000, hemoglobin 40% and leucocytes 7500. In 4 of the other cases the red 
cell-counts varied from 4,000,000 to 4,500,000. The highest count was 5,348,000. Of 
9 of these cases in but 2 was a leucocytosis present. 

Our other cases of carcinoma showed no striking features except I of the testicle, 
with 2,832,000 red cells and 9600 leucocytes. Cancers of the kidney are said to have 
usually high leucocyte counts, even 54,000, but we have seen no such case. In cancers 
of the thyroid the count may be 71,000 and in those of bone even 52,700. 

Sarcoma has much the same effect on the blood as carcinoma and some 
think a worse. We can not believe this from the study of our cases unless 
the disease involves especially the bone-marrow or the lymph-glands. In 
those cases a severe anemia, resembling a primary pernicious anemia, and 
a high leucocytosis are the rule. In Hayem's case of osteosarcoma the red 
blood cell-count was 663,400; of v. Limbeck's 2 cases, in 1 the red cell-count 
was 1,118,000, hemoglobin 28% and leucocytes 68,200; in the other the 
red cell-count was 2,240,000, hemoglobin 48% and leucocytes 54,000. Yet 
other patients with this disease have counts even above 6,000,000. Nu- 
cleated reds are said to be less numerous in sarcoma than in carcinoma. 
The hemoglobin is said to be more reduced by sarcoma than by other neo- 
plasms. The average given is about 50%. Thirty per cent, is a not rare fig- 
ure and cases with hemoglobin even below 10% have been reported. The 
leucocyte counts in cases of osteosarcoma average about 17,000; that is, 
they are higher and the picture more often resembles leukemia than in cases 
of carcinoma invading the bone. The polymorphonuclear neutrophiles are 
less increased than by carcinoma but they may be increased when there 
is no leucocytosis. In some cases with little other evidence of metastases to 
bone the eosinophile cells are greatly increased, even to 50%. Myelocytes 
are sometimes present. The old question whether some of these white cells 
in the blood may not be free sarcoma cells is often raised, for it is the small 
mononuclears which are particularly increased. 

Lues. — Lues, according to v.Limbeck, illustrates best the dictum that 
no one blood picture can be considered characteristic of any one disease. 
The blood picture in lues may simulate all other blood pictures from 
chlorosis to pernicious anemia of even the severest grade with a count of 
only 428,000 cells. Some cases of acquired lues have a practically normal 
blood, but this is unusual. 

It is important not to confuse the anemia due to the disease itself, seen 
in untreated cases, with that due to vigorous mercurial treatment. 

During the primary stage sl severe chlorotic anemia is the rule. One 
following the large European skin clinics is struck by the weight given to 



THE BLOOD 653 

this anemia, particularly in trie case of women, in the diagnosis of a primary 
sore. Some say that at first, while the count remains normal, the hemo- 
globin will diminish considerably. Later,one of the first signs of the dissemi- 
nation of the disease is the appearance of the skin rash and a further dim- 
inution of hemoglobin. The count may remain nearly normal notwith- 
standing a loss of hemoglobin of 25 to 30%. If the lues is untreated the 
hemoglobin may soon reach as low as 2 5% and the red blood-cell-count drop 
even at the rate of 23,000 cells per day. The severity of the anemia de- 
pends on the condition of the patient, his age, treatment, etc. In well- 
treated cases the regeneration of the blood is rapid. 

Leucocytes. — In an adult a high lymphocytosis and an eosinophilia 
would suggest lues. In a child this blood picture might suggest rickets also. 
A low hemoglobin per cent, and a high percentage of small mononuclears 
would indicate a severe case. 

The leucocytes in the primary stage are normal, or there is a slight 
leucocytosis with an increased percentage of lymphocytes. If mercury is 
given the percentage of the polymorphonuclear neutrophiles will rise. This 
is the reverse of the action of mercury in a normal case. 

During the secondary stage the leucocytes vary from 12,000 to 16,000, 
due to an increase of lymphocytes and eosinophiles. The latter are in- 
creased especially with the papular syphilide. 

The tertiary stage is often accompanied by a severe anemia and a leu- 
cocytosis due to a high lymphocytosis, which is of aid in excluding per- 
nicious anemia. Myelocytes are present in severe cases. 

In 19 cases of secondary lues the red cells were but slightly diminished. (The mini- 
mum count was 4,200,000. In 6 it was above 5,000,000.) The hemoglobin was more 
affected than the count of red cells. (This varied from 40 to 90%, the mean, 75%; and 
the color-index varied from 0.5 to 0.9, the mean 0.7.) The leucocytes, as a rule, were 
normal (in 11 cases below 10,000, in 3 between 10,000 and 12,000) except in 5 cases with 
high fever (luetic fever of secondary stage) in 4 of which they ranged between 12.000 
and 24,000 and dropped with the temperature. 

There was a slight rise of the leucocyte count during the primary stage 
(the average was 9000). During the secondaries the count, depending on 
the skin lesion and on the fever, varied from 9000 to 24,000 (in 1 case even 
50,000), the average from 12,000 to 15,000. During the tertiary stage the 
counts varied greatly; in some cases there was a slight rise, in others a 
leukopenia. In hereditary lues the count has been found high, from 12,000 
to 24,000. 

In hereditary and tertiary lues the red cells are seriously affected in 
number, size and color. Megaloblasts are common. The blood picture, 
especially of the long-standing cases, may resemble that of primary per- 
nicious anemia; yet, as in cancer, the megalocytes do not predominate as 
they do in pernicious anemia. Many cases of anemia in children reported 
as luetic were probably cases of anemia pseudoleukemica infantum. Miller 



654 CLINICAL DIAGNOSIS 

reported a case with 720,000 reds and 18% hemoglobin, with normoblasts, 
megaloblasts, even gigantoblasts, microcytes and poiklilocytes present. The 
anemia of the hereditary lues of infancy may be fatal. The average leuco- 
cyte count of 25 cases was 7050. Large nucleated reds containing little 
hemoglobin aid in diagnosis (Cima). 

Following mild mercurial treatment the red cells may rise even 100,000 
cells a day for about 14 days, until even a slight hypercythemia is present. 
But this rise is often preceded by a drop (Justus' test, see below) but some- 
times with a hemoglobinuria which is followed by rapid regeneration. If 
the mercurial treatment is carried too far, that is for 24 or more days, it 
may itself produce an anemia. 

Of 23 of our cases, 7 of which were of cerebral lues, the red cells were above 5,000,000 
in 9 cases and between 4,000,000 and 5,000,000 in 10. The lowest count was 2,870,000. 
The color-index varied from 0.4 to 0.9, the mean was 0.67. The leucocytes were below 
10,000 in 20 of 29 cases. In the other 9 cases they varied from 10,000 to 18,500 (this 
case had large gummata) ; 6 were cases of high luetic fever; another was of the malignant 
type (leucocytes 16,000, no fever). In 1 cerebral case the leucocytes numbered 3000, 
in another 2100. 

Justus* Test. — If a large inunction or injection of mercury is adminis- 
tered a patient with lues before the rash and yet after there is general 
glandular enlargement, and also during the secondary or tertiary stages, 
and in hereditary lues provided the disease is at the time advancing, the 
hemoglobin will at once drop from 10 to 20% and during the next few days 
will return to normal or even to above normal with improvement of all the 
symptoms. This drop, which is both rapid and considerable, is said to be 
specific for a case of active lues. It is not positive during the primary stage 
while the infection is limited to the chancre and its neighboring glands, 
therefore it cannot be used early to differentiate between a hard and a 
soft chancre. 

The explanation suggested is that the mercury destroys the red blood- 
cells which are already damaged by the disease and stimulates the produc- 
tion of new one?. 

This test is not as specific as was first claimed and yet it is valuable. 

Renal Disease. — The kidneys play an important part in the control of 
the composition of the blood, hence in nephritis the plasma changes are 
early and important: a loss of albumin, a lowered specific gravity and in 
general all the signs of a chronic secondary anemia. For the further chem- 
istry of the blood in nephritis see pages 539 and on. 

In acute hemorrhagic nephritis especially the count may fall to a 
low point, even to 1,000,000, but usually the anemia is moderate, and of this 
much is only apparent. 

In 12 recent cases of acute nephritis there were but 2 low counts, 2,600,000 and 
2,900,000; there was a leucocytosis in 5 of from 11,400 to 18,900. Of Cabot's 50 cases, 
the lowest red count was 3,568,000 but the leucocytes were above normal in 31 of these 



THE BLOOD 655 

cases. (The highest was 50,000. ) Cabot thinks the leucocy tosis due in part to hematuria 
or uremia. But since nephritis is, or is part of, an acute febrile and probably infectious 
disease a leucocy tosis is to be expected. 

In chronic nephritis many factors come into play, nearly all of which 
tend to produce an anemia. Among them are a subacute often latent pyo- 
genic infection; the disturbances of circulation; the edema and hydremia; 
the disturbances of the gastro-intestinal tract, vomiting, diarrhea, poor 
appetite, and the influence of the purges. The result is often a lowered 
count, a still more lowered hemoglobin per cent, and a hydremic plasma. 

In 103 cases of chronic nephritis the red cells were 1,700,000 in 1 case, between 
2,000,000 and 3,000,000 in 13 cases, between 3,000,000 and 4,000,000 in 25 and over 
5,000,000 in 19. The mean was 4,500,000. 

The hemoglobin in 99 cases was between 20 and 30% in 3 cases, from 30 to 50% in 
29 and above 80% in 17. The mean was 62%. The mean color-index was tnerefore 
0.7, which is about normal. 

The leucocytes in 80 cases without uremia were below 5000 in 4 cases, from 5000 to 
10,000 in 43 and above 10,000 in 33. (The highest were between 20,000 and 30,000.) 

In 33 cases with uremia the highest leucocyte count was 25,900. It was above 
10,000 in 15 cases and below 5000 in 2. The mean was about 9000. It is seen that in 
our cases the uremic syndrome was not mirrored in the blood. 

The cases of nephritis which resemble pernicious anemia form a most 
interesting group. 

The case which Dr. McCrae 154 reported is a good illustration of this. The patient 
was a man 39 years old whose red cell-count was 1,400,000, hemoglobin 27% and leuco- 
cytes, 7000 (pmn. n. 88%; s.m. 8%; l.m. 2%; and eos. 2%). There was no poikilocytosis 
and but 1 nucleated red was found. The urine was of low specific gravity and contained 
much albumin and many casts. 

Cabot reports such a case with 1,468,000 reds, hemoglobin 23% and leucocytes 3800 
(pmn. n. 70%; l.m. 4.4%; eos. 2.6%; megaloblasts, normoblasts and poikilocytes were 
present). 

In the case of Labbe the red blood-count was 500,000 and the hemoglobin 2 gms. 
The cells were pale and irregular in form and size. The nucleated reds were rather 
small. Mononuclears made up 50% of the leucocytes. Recovery was rapid. He 
suggests that anemia was for the most part apparent and due to the dilution of the 
plasma. In another case the red blood cell-count was 418,500 and the color-index over 
1; and in a third the count was 1,000,000. At autopsy in such cases nephritis was the 
only lesion found. There were in these cases practically no signs of blood destruction, 
nor of regeneration, nor of megaloblastic degeneration of the marrow. 

We mention 2 other cases of chronic nephritis with marked arteriosclerosis. One 
was awoman 54yearsof age, with red cell-count 2,800,000, hemoglobin 50%, and leuco- 
cytes 6000. The other was a man 32 years old, with red cell-count 1,772,000, hemo- 
globin 22% and leucocytes 50,000 (of which 91% were pmn. n.). The leucocytes later 
rose to 116,000. He left the hospital unimproved. 

In interstitial nephritis the count is normal at first, and sometimes re- 
mains so to the very end. The condition of the heart is in this connection 

154 Johns Hopkins Hosp. Bull., October, 1902, p. 245. 



656 CLINICAL DIAGNOSIS 

important. During the acute exacerbations of the nephritis, however, 
a slight lowering of the count is common. This is perhaps due to the hy- 
dremia. In 2 cases of bilateral cystic kidney the red cell-counts were 
1,200,000 and 2,800,000; and the leucocyte counts, 13,500 and 36,000. 

Diseases of the Liver. — Catarrhal Jaundice. — " Occasionally in 
catarrhal jaundice there is a slight leucocytosis at the onset but otherwise 
the blood is normal, although some degenerative changes of the red cells 
are met with in severe cases" (Cabot) . An increase in resistance, in rigidity 
and in size is claimed for the erythrocytes. 

Of 27 of our cases the red count was normal or even above normal in 16; the lowest 
was 3,000,000; and the mean, in the male patients, 5,000,000. An interesting feature 
was a rise in this count while in the hospital of from 300.000 to 750,000 cells. Of the 
27 cases, in 20 the leucocyte count was 10,000 or below; in 3, from 10,200 to 19,500; and 
in 4 from 14,200 to 19,500. These cases all had a slight fever. These cells fell rapidly 
to normal after admission. A leucopenia may follow in some cases (Bezancon and Labbe). 

The plasma is bile-stained. The coagulation time slow. 

Toxic Jaundice. — There were in the Johns Hopkins Hospital clinic 
3 fatal cases of toxic jaundice. In 1 the red cell-count was 3,570,000, 
hemoglobin 65% and leucocytes 11,400. In the second, these figures 
were 5,280,000, 75% and 7000; and in the third, 5,400,000, 65% and 
12,500 respectively. 

Gall-stones. — A mild leucocytosis is the rule during an attack of gall- 
stone colic; a high one is rare. In the Johns Hopkins Hospital's 36 cases 
these cells rose suddenly during the colic to about 15,000, but in the cases 
of stone in the common duct with chills and fever, they rose even to 24,700. 
The red cell-count varied from 2,800,000 to 6,400,000, the mean was 
4,300,000. In a case with hemorrhage they fell to 1 ,880,000, the hemoglobin 
to 23%, while the leucocyte count was 17,500. The coagulation time of the 
blood of a jaundiced patient is often lengthened and so should be tested 
before any surgical operation which should be postponed until, as the re^ 
suit of therapy, this becomes normal. 

Cholecystitis. — In cholecystitis the leucocyte count is invariably high, 
from 20,000 to 27,000 (Bloodgood). In 1 of our cases it was 46,500. As 
the case becomes chronic the count falls nearly or quite to normal. 

Cholangitis. — Of 5 cases of cholangitis the leucocytes were 16,000, 
33,160 (fatal), 15,600 (t.° 103. 5 ), 9000 (t.° 103 °) and 6,400 (t.° 106 ; fatal). 

Abscess of Liver. — In cases of abscess of the liver the leucocyte counts 
are high while the temperature is high, but are lower or normal when the 
temperature is normal. Futcher 155 found the average in 15 cases to be 
18,350 and the maximum 53,000. The red cells in these cases varied from 
2,600,000 to 5,600,000, the mean, 4,200,000; the mean of hemoglobin 
was 60%. 

155 Jour. Am. Med. Assoc, August 22, 1903. 



THE BLOOD 657 

Cirrhosis of the Liver (Atrophic). — Early in cases of cirrhosis of the 
liver the red cell-count is normal while later an anemia may develop. 
Da Costa's average was 3,404,000 and Cabot's, 3,580,000. In 1 case it fell 
to 1,300,000. The leucocyte counts are normal or low. 

In the Johns Hopkins' 32 cases the red cells varied from 3,100,000 to 5,900,000, 
the mean 4,500,000. The hemoglobin mean was 68%. The leucocyte counts in 30% 
of our cases were over 10,000 and the highest was 16,000. 

Hypertrophic (Hanot's) Cirrhosis. — Hayem reported a case with 
extreme anemia. 

In the Johns Hopkins' series there were 5 cases. In 2 the count was high, 7,800,000 
and 8,500,000, and in 1 as low as Hayem's case — 1,504,000. In this last case the hemo- 
globin was 28% and the leucocyte count 6100. (This count rose later.) In 2 there was 
a leucocytosis (11,000 and 12,800). 

Acute Yellow Atrophy. — In the cases of acute yellow atrophy thus 
far reported the red cell-counts were normal and the leucocyte counts slightly 
elevated. In one case of the Johns Hopkins' clinic, a boy 14 years old, the 
red count was 4,800,000 and the leucocyte count 12,700. 

Leprosy (v. Limbeck). — In leprosy the blood may for years be normal 
but later a pseudochlorosis develops with a normal leucocyte count. After 
general malnutrition begins the anemia becomes more marked and yet is 
rarely very severe. In 1 case, however, the red cell-count was 2,290,000 
and the hemoglobin 55%. The leucocyte counts have varied from 4000 to 
8000 per c.mm. 

Heart Disease. — While cardiac compensation is good the blood is 
normal, but with acute dilatation and fall of the blood-pressure the blood 
becomes hydremic, hence the count falls. Later with chronic passive con- 
gestion and cyanosis the red cell-count rises and so may conceal an anemia. 
The most marked anemia is seen in aortic valvular insufficiency, as in 1 
case with red cell-count 3,400,000, hemoglobin 30% and leucocytes 8000. 
If, as the result of proper therapy, the anemia improves this may aid the 
heart to regain its compensation. In congenital heart disease with extreme 
cyanosis the picture is particularly interesting since there often is a poly- 
cythemia, the red cell-count between 8,000,000 and 9,000,000. 

During the loss of compensation in 29 males with pure mitral disease the count 
varied from 3,000,000 to 7,500,000, the mean 6,200,000. In 46 women the mean was 
4,700,000, but the extremes were 3,500,000 and 8,000,000. One noted interesting jumps 
of from 1,000,000 to 2,000,000 cells while these cases were under treatment. 

In 37 cases of pure aortic disease the mean red cell-count was 5,200,000. These 
cases as a rule showed a lower count on each successive admission. 

In 29 cases of arteriosclerosis (no important cardiac lesions) the mean was 5,200,000. 
In 34 cases of aneurism of the thoracic aorta the mean was 5,500,000 and in 5 men with 
aneurism of the abdominal aorta it was 4,500,000 

42 



658 CLINICAL DIAGNOSIS 

Addison's Disease. — A hypocythemia is the rule in Addison's disease, 
the red cell-counts varying from 2,000,000 to 3,000,000. In 1 case it was 
1 , 1 20,000. In other cases, however, the count may be even above 7 ,000,000. 
Some consider that any anemia in this disease is due to a complication; 
others, that there is always an anemia but that this is covered by a con- 
centration of the blood, and cite a case with true oligemia and a count 
of 4,774,000 cells. 

Myxedema. — In myxedema the red cell-count may be normal, above 
normal, or diminished. Many report an anemia which improves with treat- 
ment. Some report that the diameter of the red blood-cells is increased 
and also the presence of many nucleated reds (that is, an infantile condition 
of the blood) . The platelets in a recent case were much increased. 

Rickets. — Anemia, generally of a mild grade, is the rule in rickets, but 
sometimes it becomes severe, rapid in its development and even pernicious. 

Scurvy. — The red cell-counts reported in scurvy have varied from about 
3,000,000 to 4,000,000 cells. The cases with many hemorrhages have a 
much more intense anemia. In Buchard's case, e. g., after 3 weeks with 
considerable epistaxis, the count was 557,000. In some grave cases macro- 
cytes, microcytes and fragmented reds have been found. The color-index 
is reported low. 

The Blood in Inanition. — The effects on the blood of a healthy man of 
a 3 1 -day fast were studied by Ash l56 who found a very slight actual loss in 
hemoglobin, more marked during second 10 days (of but about 7%), mod- 
erate fluctuations in the water content of the blood, particularly during 
first half of the period, a decided rise in polymorphoneutrophiles in the early 
days (to 79%) and an increase in coagulability, especially after the first 2 
weeks (coagulation time on the 21st day was 55 seconds). He found the 
blood as a whole distinctly resistant to the effects of uncomplicated inanition. 

THE VALUE OF BLOOD EXAMINATION 

By the examination of the fresh blood the diagnosis of some cases is made 
or is suspected. Among these are malaria, especially the forms without defi- 
nite paroxysms, which often pass as typhoid fever, meningitis, uremic 
coma, pernicious anemia, appendicitis, tuberculosis, dysentery and even 
Raynaud's disease (cases with superficial gangrene). The failure to look 
at a smear of blood may in these cases result in the unnecessary death of a 
patient. And yet not every patient with malarial organisms in his blood 
should then be treated for malaria. He may be a chronic carrier who now 
has typhoid fever, etc. Trypanosomiasis can be recognized by the blood 
and the Leishman- Donovan bodies by splenic puncture. Pernicious 
anemia is often overlooked although a cursory glance at a fresh blood 
specimen would save some patients from a course of treatment for jaundice, 

156 Arch, of Int. Med., July, 1914, xiv, p. 8. 



THE BLOOD 659 

peripheral neuritis, or tabes. The practical value of 3 minutes spent making 
and glancing at a fresh blood specimen is shown by the fact that the 
majority of our cases of myelogenous leukemia come to the surgical side 
for "abdominal tumor." The diagnosis of lymphatic leukemia, acute 
leukemia and pseudo leukemia can be made only in this way. 

For the early diagnosis of typhoid fever, measles and influenza a leu- 
copenia is valuable, while a leucocytosis would speak in favor of pneu- 
monia, scarlet fever, acute epidemic cerebrospinal meningitis and various 
abscess formations, as of the liver or brain, etc. Early in typhoid fever a 
blood culture often settles the question, later a Widal test might do so. 

The presence of a leucocytosis is very valuable in the diagnosis of 
pneumonia, especially central, and that of children and drunkards. In an 
ever-increasing number of cases of trichinosis the diagnosis has been 
suggested by the eosinophilia alone while the blood examination has the 
same value in cases of chronic poisoning with coal-tar products notwith- 
standing the denials of the patients. Various tuberculous infections are 
thus differentiated, also the secondary anemias due to cancer from pri- 
mary anemia, etc. 

For the surgeon, blood examination is usually synonymous with leu- 
cocyte counting. The question he usually asks is, "Should I operate or 
not ? " on a doubtful case of appendicitis, typhoid perforation, etc. In 
these cases the leucocyte curve should be determined rather than a 
single count. 

Both the medical man and the surgeon should remember that 1 count 
is seldom enough, any more than is 1 temperature reading. It is the curve 
that has value. 

For American students the message is, less routine blood-work but a 
better quality of that which is done. The examination of the fresh specimen 
will save a great many unnecessary routine counts. Also interpret your 
findings in terms of the individual case, for changes in blood cells or serum 
are symptoms to be translated, not signs to be obeyed. 

MALARIA 

The following are a few of the terms used in this chapter which may need definition. 
Schizogone, the asexual generation; schizont, or monont, an individual organism belonging 
to the asexual generation; merozoite, a segment, i.e., a hyaline; garnet oschizont, the sexual 
generation; gamete form, I organism of the sexual generation. Of the gamete forms, the 
macrogamete is the female cell; the microgametocyte, the parent male cell, and the micro- 
gamete is the male cell, i.e., is I " flagellum " of the microgametocyte. Sporogone, the 
cycle in the mosquito; vermiculus or ookinet, the motile fertilized macrogamete; zygote, 
oocyst, sporoblast, are terms given to the spore cysts; sporozoit, the young sexual form 
which develops in the sporoblast and which, when inoculated into the human blood, 
becomes a hyaline. 

By malarial pigment is always meant the transformed hemoglobin, the 
brown granules of which are seen in the fresh specimen. This term is never used of 
chromatin granules. 



660 CLINICAL DIAGNOSIS 

Hyaline always means a non-pigment ed young form. A ring form 
is the shape which any hyaline may assume; it is not a "kind " of organism. 
Presegmenters are full-grown parasites before segmentation has appeared, 
the pigment of which ha,s accumulated into i or a few masses. The term 
granule is used in several senses when describing malarial blood. It may 
mean a particle of malarial pigment, or a small mass of chromatin, or a 
degenerated area of the red blood-cell (Plehn's granules). 

The examination of the fresh malarial blood is often more satisfactory 
than is that of the stained specimens since for the diagnosis of the various 
forms of parasite the color of the red cell, the refractivity of the parasite's 
protoplasm, the rapidity of its ameboid motion and the speed of vibration 
of its pigment, all are important. It also is true that one is less likely to 
be deceived by artifacts in fresh than in stained blood. On the other hand 
the parasites are easily found in stained specimens and, when very few, 
the Ross method alone may make diagnosis possible, 

The Organism of Tertian Fever; Hemameba Vivax (Grassi); Plas- 
modium Vivax (Plate III). — This is the commonest form of malaria in 
this country. Since the cycle of development of this parasite requires 
approximately 48 hours the paroxysms in the case of a single infection will 
occur on alternate days, granting that the infection is intense enough to 
cause paroxysms. In case there is a double infection there will be a 
paroxysm each day, "quotidian " fever, and the 2 groups will be seen in 
the blood. Three groups very rarely occur, but we have seen 2 or 3 cases. 
The grouping of these organisms is fairly definite, i. e., all of the parasites 
of 1 group develop quite in unison. The paroxysms occur during segmen- 
tation and last from 12 to 14 hours. The hyaline form of the tertian para- 
site (2-4) is a little over 2ja in diameter, is colorless, non-pigmented and often 
disk-shaped, with an undulating periphery. It is either on or in a cell which 
even now may be a trifle swollen. It now makes very rapid ameboid move- 
ments, which produce an extraordinary series of changes of shape and 
position and which at any time may assume the typical ring form 
once supposed to be characteristic of the asstivo-autumnal parasite. 
This ring is usually a little thicker at one point, due to the mass of chro- 
matin, hence the name "signet ring form." In one red cell may be 1, 2 or 
even 5 such hyaline forms. In about 12 hours the infected corpuscle (6-7) 
will be a little larger, a little paler, but still has a sharp, smooth, round 
margin. The organism is exceedingly ameboid. The pseudopods often are 
numerous and so thread-like and pale that their connections can scarcely 
be seen and so the cell seems to contain a number of disconnected globules of 
pigmented protoplasm. The protoplasm is so little refractive that the out- 
line of the parasite is difficult to make out. The malarial pigment is now 
present. It is moderate in amount, massed at the ends of the pseudopods 
and consists of very fine, light-brown granules, which dance with a motion 
so rapid that waves in the protoplasm must be assumed to explain it. At 



PLATE in 

The Parasite of Tertian Fever. (Drawn by Mr. Brodel for Thayer and Hewetson's paper, The 
Malarial Fevers of Baltimore, Johns Hopkins Hospital Reports, Volume V.) 
I. Normal red corpuscle. 
2, 3, 4. Young hyaline forms. In 4, a corpuscle contains three distinct parasites. 
5, 21. Beginning of pigmentation. The parasite was observed to form a true ring by the confluence 
of two pseudopodia. During observation the body burst from the corpuscle, which became 
decolorized and disappeared from view. The parasite became, almost immediately, 
deformed and motionless, as shown in Fig. 21. 
6, 7. 8. Partly developed pigmented forms. 

9. Full grown body. 
10-14. Segmenting bodies. 

15. Form simulating a segmenting body. The significance of these forms, several of which have 
been observed, is not clear to the writers, who have never met with similar bodies in 
stained specimens so as to be able to study the structure of the individual segments. 
16, 17. Precocious segmentation. » 

18. Macrogamete. 
19, 20. Fragmenting extra-cellular bodies. 

22. Microgametocyte. 
23, 24. Vacuolization. 

The Parasite of Quartan Fever. 

25. Normal red corpuscle. 

26. Young hyaline form. 

27-34. Gradual development of the intra-corpuscular bodies. 

35. Full grown body. The substance of the red corpuscle is no more visible in the fresh specimen. 
30-39- Segmenting bodies. 

40. Female gamete. 

41. Microgametocyte. 

42. Vacuolization. 



The Parasite of Terhan Fever 



PLATE III. 






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THE BLOOD 661 

the end of 24 hours the cell (Plate III, 8) is somewhat larger and paler but 
still round in outline. The organism now fills about % of cne cell. That is, 
it has lived }4 its cycle and yet has attained but % of its adult size. It is 
still ameboid, but less actively so. The pigment has increased in amount, 
is a little darker, a little coarser, a little quieter and is now evenly 
distributed through the substance of the parasite. The nucleus of these 
forms sometimes can be seen in the fresh specimen as a globular body at 
the end of a pseudopod. 

During the latter half of the cycle the growth of the parasite is much more 
rapid. At 40 hours the parasite (9) is practically full-grown. The red cell 
is now about i}4 times its normal size and so pale that its outline will hardly 
be seen. The organism is from 8 to 10/z in diameter, is round and so little 
refractive that it is practically impossible to distinguish between parasite 
and corpuscle. The pigment is more abundant and is evenly dis- 
tributed throughout the parasite, an important point in diagnosis since 
in the quartan at this age it will be practically all in the periphery, 
and in the asstivo-autumnal at the center. 

The next stage is the "presegmenter." The corpuscle is now almost or 
quite invisible. The pigment granules move in regular lines to collect in 1 
or more irregular clumps. The organism is now a "segmenter " (10-17). 
It becomes slightly more opaque, more refractive, and the corpuscle is no 
longer seen. Retractile dots, from 15 to 20 in number, appear irregularly 
in the body of the protoplasm, crenations are seen at the margin and lines 
of separation appear around these refractive dots marking off the future 
segments. The segments now become more sharply defined until finally the 
parasite is a. clump of 15 or 20 discrete circular masses with a refractive dot 
in the center of each, irregularly arranged or forming 2 quite concentric 
circles. The pigment is left in masses between these segments. The 
segmenter now seems to burst since the young organisms suddenly spring 
apart. Each segment is a hyaline ready for a new cell as host. 

The whole cycle may occur in the peripheral blood, but the number of 
segmenters found will not be as large as would be expected from the number 
of parasites seen a few hours previously, since so many of them have accu- 
mulated in the internal organs. A few hours after the first segmenters 
appear the chill begins. 

The above is a description of a typical tertian parasite. One finds, how- 
ever, forms which differ somewhat. In one, the parasite develops more pig- 
ment, in large coarse granules of a lighter brown color than that of the 
quartan or the adult asstivo-autumnal and these form dense clusters at the 
ends of the pseudopods filling them so completely that the granules cannot 
dance at all. The fine thread-like pseudopods of these parasites stand out 
with great distinctness. The infected cell is often not much swollen but 
is very pale. In one such case, however, all the full-grown forms found were 



662 CLINICAL DIAGNOSIS 

in cells from 8.5 to 13.3 11 in diameter. Pigmented leucocytes are common 
(perhaps since the pigment granules are so conspicuous) . 

The grouping of this form does not seem to be as definite as that of the 
typical tertian and so the chills are slightly longer than usual. 

Extracellular Tertian Forms. — It is not at all uncommon for the 
tertian parasites to burst from their cells and die (19-21). These degen- 
erated forms may, a short time after the specimen is made, be the only ones 
seen. The organism is often seen to "run out" of the red cells as if through 
a very fine hole, leaving but a shadow of the cell. After the parasite is 
free in the plasma the pigment gradually becomes quiet, the protoplasm may 
then break up into fragments, forming a string of 4 or 5 small pigmented 
spherical masses (20-21), or it may become deformed, swollen and vacu- 
olated, the formerly so-called "sporulating forms" (23, 24). 

The tertian gametocytes would appear in fresh specimens to be full 
grown forms, often extracellulars, but when stained the shell of the cor- 
puscle may be seen. Like the crescents of the asstivo -autumnal parasite to 
which these correspond, they can be found in the blood at all times after the 
infection has continued for a few days. The macrogamete (18) formerly was 
considered a cadaveric form, and was known as a ''swollen extracellular." 
These large organisms, some 3 or 4 times the size of a red blood cell, have 
abundant pigment often in the form of coarse rods which are in very active 
movement. Their nucleus is about 3.5M in diameter and is often evident 
in the fresh specimen as the only portion of the parasite which the pigment 
granules do not invade. The extreme vitality of these cells is astonishing 
(as might be expected from the fact that it is their function to continue the 
life of the organism in the mosquito) for the pigment will dance actively for 
even after 18 hours if the microscope is kept in a warm room. The micro- 
gametocytes are from 8 to iom in diameter. When studied on the warm 
stage their pigment, at first in active motion, soon forms a circle around the 
center. Then, as though stirred up by something within the cell, the margin 
may undulate and the "flagella" (erroneously so called), 4 or 5 in number, 
burst out. These ' ' flagella ' ' are the microgametes or male elements. They 
are threads of chromatin from 2 to 3 times the diameter of a red blood-cell 
in length and often rendered irregular by fusiform masses of protoplasm 
containing pigment granules. These microgametes break loose and thrash 
their way for more than an hour among the red cells leaving behind only 
a small cell with its pigment near the center. This "flagellation" is seen 
first in from 15 to 20 minutes after the blood has been drawn which is proof 
that it does'not occur in the body, but under the stimulus probably of the 
lowered temperature. 

Hemameba Malariae, the Parasite of Quartan Malaria (Plate IV). — 
Of this rare form we see few cases now although in some localities it is at 
times the predominant organism. Since its cycle of development re- 
quires 7 2 hours, if but one group is present there will be one paroxysm on 



THE BLOOD 663 

each fourth day; if 2 groups, there will be 2 days with paroxysms and 
then a free day, followed by 2 more paroxysms, etc.; if 3, the patient will 
have quotidian fever, providing each group is large enough numerically 
to cause a chill. The grouping of this parasite is even more uniform than 
is that of the tertian ; that is, the forms of the same group keep more nearly 
of the same age and hence the paroxysms are slightly shorter, often requir- 
ing but 10 hours. 

. The quartan hyalines (26) cannot be distinguished from the tertian 
parasite, but a little later, when the pigment appears (27), the granules 
are seen to be coarser, blacker and less actively vibratory than the tertian. 
As the parasite grows the infected cell becomes smaller, shrunken and its 
margin irregularly crenated, but much deformity is rare. The protoplasm 
of the parasite is very refractive so it can easily be seen. This parasite is 
only sluggishly ameboid. 

In 24 hours the infected cell is still smaller, is crenated and brassy in 
color. The organism is very distinctly made out since so refractive, is 
oval and sluggishly ameboid. The pigment is coarse, blackish-brown in 
color, is clustered at the periphery, especially on one side, and is quiet. 
The parasite soon fills from one- third to one-half of the cell (30, 31), 
becomes rounder, and is no longer ameboid. The infected cell may be 
more shrunken, more crenated and more brassy, although some seem 
but little altered. The coarse, black pigment is entirely at the periphery. 

During the third day only a rim of the red cell is left and this is usually 
of a dark, brassy color. The organism (32-34) is now full-grown and aver- 
ages, most authorities say, about 7 /jl in diameter. 

Of 135 full-grown quartan parasites, some of them segmenters, which we have meas- 
ured, 60% were from 7.4 to 8.iju in diameter, and only 18% from 6.2 to 7/x. Of those 
% grown, 43% were in cells from 6.2 to 7ju in diameter, the rest in cells of normal size. 

At 60 hours the red cell is scarcely visible. The organism is round 
or elliptical and motionless (35). The coarse dark pigment is all at 
the periphery. 

The pigment now flows to the center in definite streams along definite 
radial channels, thus giving a beautiful wheel-like picture (36), and finally 
collects in a clump at the center. These are the presegmenters. The seg- 
menters present an attractive picture. The organism becomes very waxy 
in appearance; a single row of refractive dots form a circle at the periphery 
which becomes crenated around each and lines of division start from these 
crenations and run to the center, forming from 6 to 1 2 rays like the petals 
of a flower, hence the names "daisy," "Marguerite," or "rosette" form 
(37, 38). These segments then separate as in the tertian. The quartan 
lives its full cycle in the peripheral blood, hence the number of parasites in 
the peripheral blood does not diminish as the parasite grows older. 

The gamete forms (40, 41) are seldom seen. They are similar to but 



664 CLINICAL DIAGNOSIS 

somewhat smaller than those of the tertian. Flagellation occurs in the 
same way. The extracellular degenerated forms are found, although the 
parasite keeps within the cell much better than does the tertian. 

^stivo-autumnalFever;Plasmodium Precox; Hematozoon Falciparum 

(Plate IV) . — iEstivo-autumnal fever is a common form of malaria partic- 
ularly in the Tropics and is the most dangerous of the three. In fresh in- 
fection the grouping of the organisms is quite definite and the fever definitely 
intermittent, but soon the members of a group lose their unison of growth 
and parasites of all ages may be found in the same blood (and spleen) as a 
result of which the fever becomes more and more continuous. The duration 
of the cycle is rather uncertain. Doctor Thayer considers that while usually 
it is about 48 hours, it may vary from 24 to perhaps 72. 

All students, it is said, pass through the stage of desiring to divide this 
form into "benign," a "malignant," or "pigmented," "non-pigmented," 
etc., varieties, but most recover, especially those who follow the splenic 
blood carefully. In this locality we are often impressed by the great dif- 
ferences seen in the asstivo-autumnal parasite, especially as regards the 
amount of pigment it develops and the number of adult forms which 
appear in the circulation; but no division of this group has stood the 
test of time. 

The hyalines of the aestivo-autumnal are similar to those of the tertian 
and the quartan except that they are perhaps slightly smaller and are 
more apt to assume the signet -ring form. They are very refractive, 
hence easily seen, but may at any time lose this refractivity and become 
ameboid exactly as does the tertian. In a severe infection even five rings 
may occupy one cell. 

As the parasite (7-12) grows a very slight amount, usually but 1 or 2 
granules, of pigment appears. These are so fine that they are easily over- 
looked, are motionless as a rule, although sometimes they dance 
slightly and for the most part are seen at its periphery. The infected red 
cell is usually much shrunken, crenated and brassy, even when the parasite 
is young, and yet some infected cells appear normal. Many red cells 
which contain no parasite also show similar changes. The parasite at 
this stage fills about }{ of the cell. As a rule the infected cells now disap- 
pear from the peripheral circulation and to study their further develop- 
ment the spleen must be punctured . Their departure is so sudden that 
in 2 hours a large brood may disappear. Rarely are older parasites 
found in the peripheral blood (Fig. 117). In these cases and in the 
blood obtained by splenic puncture, the pigment increases considerably 
in amount as coarse, dark granules, although some produce practically 
none. The former may easily be mistaken for quartan forms. The 
more malignant the parasite the fewer older forms are seen in the 
peripheral blood (although in pernicious cases there is always an abund- 
ance of young parasites in the peripheral blood) and, according to 




PLATE IV 

The Parasite of ^Estivo Autumnal Fever. (Drawn by Mr. Brodel for Thayer and Hewetson's 
paper, The Malarial Fevers of Baltimore, Johns Hopkins Hospital Reports, Vol. V.) 
I, 2. Small refractive ring-like bodies. 

Larger disc-like and ameboid forms. 

Ring-like body with a few pigment granules in a brassy, shrunken corpuscle. 
Similar pigmented bodies. 
Ameboid body with pigment*- 

Body with a central clump of pigment in a corpuscle, showing a retraction of the haemo- 
globin-containing substance about the parasite. 
Larger bodies with central pigment clumps or blocks. 

Segmenting bodies from the spleen. Figs. 24-27 represent one body where the entire 
process of segmentation was observed. The segments, eighteen, in number were 
accurately counted before separation as in Fig. 28. The sudden separation of the 
segments, occurring as though some retaining membrane were ruptured, was observed. 
20-37. Crescents and ovoid bodies. Figs. 34 and 35 represent one body which was seen to 
extrude slowly and, later, to withdraw two rounded protrusions. 
38, 39, 42. Round bodies. 

40. "Gemmation," fragmentation. 

41. Vacuolization of a crescent. 
43-44. Flagellation. The figures represent one organism. The blood was taken from the ear 

at 4.15 P.M.; at 4.17 the body was as represented in Fig. 43- At 4.27 the flagella 
appeared; at 4.33 two of the flagella has already broken away from the mother body 
45-49. Phagocytosis. Traced by Dr. Oppenheimer with the camera lucida. 







3-6. 






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9, 


10, 


J 2. 
II. 
13. 




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21- 


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PLATE IV 




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THE BLOOD 665 

some, the less pigment is formed. In some cases the hemoglobin seems 
to gather around the parasite leaving an almost colorless ring of the in- 
fected cell at the periphery (13). In the internal organs the cycle seems to 
develop inside of large macrophages. The full-grown parasite is very char- 
acteristic in appearance. It is about y 2 the size of the cell (5/*), its proto- 
plasm is waxy and its pigment is all in the center (15-20), never diffusely 
scattered and never at the periphery. The segmenters vary in size from 
2.5 to 5m in diameter. The process of segmentation (21-24) resembles that 
of the tertian in that the organism breaks up irregularly into 15 or 16 very 
small segments. Very few degenerated extracellulars are found. 

Crescents and Ovoids. — The gamete forms of the sestivo-autumnal 
parasite have the same significance as those of tertian and quartan malaria 
but differ in that they have the perfectly characteristic and easily recogniz- 
able shapes of crescents and ovoids. These are found in the internal organs 
after about the fifth day of a fresh infection and appear in the peripheral 
blood on about the seventh day. The crescents (29) are slightly longer 
than the diameter of the red blood-cells, ate very refractive, have a double 
contour and usually are surrounded by a fringe of the shrunken red cell 
which in the concavity forms the so-called "bib." The pigment, in coarse 
and usually rod-shaped granules, is considerable in amount and is clustered 
at the center of the crescent either as a confused mass, a sheaf, or a ring. 
While watching the gamete it may lose its crescentic shape and become, 
first an oval (ovoids, 30-33), then a circular form (34-36), or it may resume 
the crescentic shape. Around the circular no trace of the corpuscle is left 
and its protoplasm is much less refractive than that of the crescent. Two 
forms of the circulars have been described in the fresh blood, the macroga- 
mete and the microgametocyte (see page 662). The latter may flagellate 
and fertilize a macrogamete. 157 

The phagocytes of the blood can be well studied in this form of malaria. 
In fact, the discovery of a pigmented leucocyte is almost as valuable as that 
of the parasite itself. These are the large mononuclears especially, also 
the polymorphonuclear neutrophiles and macrophages not usual in the 
peripheral blood. (See page 507 and Fig. 117). These phagocytes contain free 
pigment granules, or masses of pigment, or parasites, especially segmenters 
and flagellates. In the tertian and quartan they appear just after a chill, 
but in the asstivo-autumnal, at any or all times. The large macrophages, 
some of which are necrotic, are seen only in severe cases. These may con- 
tain organisms which are still within red cells. 

The malarial pigment is now considered to be hematin. Formerly 
it was supposed to be iron-free melanin. 

The Cycle within the Mosquito. — The cycle within the mosquito has 
been followed by several observers in the case of Plasmodium precox. 

157 See Johns Hopkins Hosp. Bull., Nov., 1897, and Oct., 1902. 



666 CLINICAL DIAGNOSIS 

The crescents in the blood which the mosquito has ingested become cir- 
cular forms. The male circulars flagellate probably in response to the same 
stimulus as under the microscope, i. e., the lowered temperature, and fer- 
tilize the female circulars. This occurs in from i to 1.5 hours after the 
mosquito has bitten. During the flagellation of the microgametocyte the 
macrogamete ripens by casting off karyosomes, polar bodies consisting of 
chromatin, and projects a slight mound through which the microgamete has 
been seen to enter. The nuclear material of the macrogamete and the 
microgamete then unite, the new fertilized cell assumes a motile spindle 
form called the " vermiculus' ' which varies in size from 20/x up, and is found 
in from 40 to 48 hours after the blood has been ingested. This vermiculus 
actively bores its way through the epithelium of the intestine and becomes 
encysted between that and the elastic layer, (Fig. 135) the "tunica elastico- 
muscularis," which forms the membrane of the oocyst. Its nucleus now 
divides rapidly (Fig. 136). This oocyst increases in size, bulging 
away from the intestinal wall until it forms a pendulous tumor in 
the body cavity (see Fig. 135) which varies in diameter from 4.5 to 30/x 
or even go/x in diameter. This stage is called the " medium zygote," 
or the "medium sporoblast," and is conspicuous because of the amount of 
pigment it contains. There may be 200 such tumors attached to the in- 
testine of the mosquito. The protoplasm now gathers itself around the 
divided nuclei (Fig. 136), a process analagous to the sporoblast formation 
of the coccydia except that here the separation is less perfect. It is now 
known as a ' ' large zygote ,"ora" large sporoblast . ' ' The nucleus of each of 
these divisions now divides into great numbers (b, c) of daughter nuclei 
which remain on the surface of the various daughter cysts. The protoplasm 
collects around each, first forming spherical cells, and these then elongate 
into threads lying parallel in masses over the residue of the sporoblast s. 
These threads are called "sporozoits." Their nuclei also become elon- 
gated. The final length of these sporozoits is about 14/* and their width 
about i//. Their protoplasm is thick, homogeneous and very refractive. 
All of the sporozoits of one oocyst ripen simultaneously. Some oocysts 
contain even 10,000 sporozoits while others contain but a few hundred. 
When ripe, the oocyst bursts into the body cavity and the free sporozoits, 
moving with a bending, gliding movement as if directed by some positive 
chemotactic influence, finally collect in the salivary glands. Inoculated by 
the mosquito's bite into the blood-vessel of men, they attach themselves 
to, and finally penetrate into, the red blood-cells, a process actually observed 
by Schaudinn. They are said to stay for some time on the surface of the 
cell before penetrating it, and it is said that if quinine is now given they will 
drop off from the corpuscle. As a rule the first chill comes on about the 
eighth or twelfth day after the mosquito bite, yet this will depend on the 
number and the virulence of the parasites introduced into the circulation. 
Since some mosquitoes contain fully 200 of these oocysts (of course not 



PLATE V 

The Blood in Tertian Malaria. 

i. A hyaline form. 

2. A young tertian, perhaps twelve hours old, with beginning granulation. 

3i 4, 5, 6. Half grown, and slightly older, forms. Three is a large cell containing two parasites 

7. A form almost full grown. 

8, 9. Full grown parasites showing division of the chromatin preceding that of the cell. 

io. A small tertian parasite in a red cell showing "stippling" (Plehn's granules). 

II, 12, 13, 14. Gamete (sexual) forms. 13, macrogamete in a cell with Plehn's granules. 

The Blood in Quartan Malaria. 

15. A very young quartan parasite. 

16. A full grown form with the chromatin still in one clump. 

17. A full grown form with the chromatin scattered. 

18. A segmenting parasite. 

The Blood in ^Estivo-Atjtumnal Malaria. 

1 a. One field exactly reproduced from the blood of a case of pernicious malaria. 

20. ^Estivo-Autumnal hyalines showing the projection of the chromatin masses from 

the cells. 

21. Blood platelets, 

22. ^Estivo- Autumnal hyalines. 

23, 24. Red cells containing more than one hyaline. 

25. A full grown aestivo-autumnal parasite. 

26. Hyalines free in the plasma. 

27. A blood platelet lying on a red corpuscle. 
28, 29. Crescents. 



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QUARTAN MALARIA. 



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EXACT REPRODUCTIOh 
OF ONE FIELD. 



^ESTIVO-AUTUMNAL MALARIA. 



STAINED WITH HASTING'S MODIFICATION OF 
ROMANOWSKI'S STAIN. ALL DRAWN TO SAME SCALE. 



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F. S. Lockteood. 



THE BLOOD 667 

necessarily all of the same age) and since some of these contain 10,000 
or more sporozoits the number of the hyalines which may be injected by 
one bite may be considerable. 

For the tertian the optimum temperature for this cycle is 2 8° to 3o°C, 
and the time 8 days ; below 1 7 to 20 C. there is no development. The quar- 
tan form can develop at a slightly lower temperature. 

The Anopheles group of mosquitoes is the only one as yet proven to be the 
host of the malarial organism. For a full description of these insects the 
reader is referred to Stevens and Christophers, and 158 Nuttall and Shipley. 159 

The Anopheles genus may be easily recognized by its "awl-shaped" 
attitude on a wall, since (see Fig. 137) its body forms a straight line with 
thorax, head and proboscis, which line forms an angle with the wall, while 
Culex (b) sits "hunch-backed," its body parallel to the wall and its pro- 
boscis at an angle of 45 degrees with its body. The genera are separated by 
the relative length of their probosces and palpi (see Fig. 138). Those of 
the Anopheles female are of equal length and scaled, while of the Culex, 
Stegomyia and Temiorhyncus females the palpi are short and insignificant. 
It is only the female Anopheles which bites. Her wings are spotted as a rule 
and she holds her hind pair of legs stretched out and oscillating in the air. 

The Anopheles egg and larva are characteristic: the former, from its 
boat-like shape and lateral air-cell floats ; the latter, from its attitude in the 
water, lying parallel with and just below the surface. 

Stained Specimens of Malarial Blood. — Very thin smears of blood may 
be stained (see page 462) by any one of the various polychrome methylene- 
blue-eosin mixtures (see page 467) . The fresher the smear when stained the 
better the preparation. 

Ross has described a method which is of great value when but few parasites are 
present. A large drop of blood is spread on the slide over an area equal in size to a I o- 
cent piece. It is then dried thoroughly in the air. The slide is then covered with water 
and the hemoglobin laked off. Care should be taken not to wash too vigorously else all 
the fresh blood will be washed off. The specimen is then stained in the usual manner. 
Mayne 16 ° has modified this method as follows : 

The thick smear is made by " dragging " the drops of blood sharply on the slide. 
Such a smear is more uniform, dries more quickly and stains more uniformly. The 
smear is dried in the air, then dehemoglobinized by covering it with 2 to 3% hydrochloric 
acid in ordinary grain alcohol until all trace of red is removed and the smear is of a clear 
white color. The acid alcohol is then washed out in several changes of water or m a 
gentle stream of running water and the specimen stained immediately. 

Tertian (Plate V, 1-14). — The youngest hyalines consist of a mass of 
blue protoplasm and a clump of carmine- violet stained chromatin. Soon 
the "achromatic zone" appears which is the "vesicular part" of the nucleus 
and this may make up the larger part of the parasite. The blue protoplasm 

158 " Malaria of the Tropics," 1905. 

159 Jour, of Hygiene, vol. i, Nos. 1 and 4; vol. iii, No. 2. 

160 Public Health Reports, Apr. 29, 1919, vol. 34, p. 837. 



668 CLINICAL DIAGNOSIS 

now often forms a wide crescentric ring surrounding this, with the chro- 
matin mass between the tips of its horns but often not quite touched by 
them. A ' ' milk-white' ' zone of Gautier often surrounds the chromatin mass 
but is not present at all ages nor in all of the same age. To just what part 
of this structure the term "nucleus" shall be applied is disputed. Stevens 
and Christophers, as well as many previous observers, use the term for the 
chromatin mass alone, while others include the chromatin mass and milk- 
white zone and others include also the much larger achromatic zone. The 
nucleus of this parasite is not specialized but rudimentary, the nuclear ma- 
terial being scattered in the cell or collected in i or more masses. 

At this point should be emphasized the necessity of studying only those 
specimens so well stained that the blue protoplasm and the red chro- 
matin can be clearly seen since so many structures can resemble a hyaline ; 
as for instance, certain degenerations of the red blood-cells and, particularly, 
platelets resting on the cells (Plate V, 21, 27). The platelets may have 
granules resembling chromatin. A platelet on a cell is always surrounded by 
a colorless zone, while the hemoglobin comes in direct contact with the 
protoplasm of a parasite; good evidence, says Ross, that the hyaline is 
intracellular and not adherent to the surface of the cell. (Argutinsky, 
Stevens and Christophers and others) . 

At the end of 24 hours the achromatic zone is a little larger, but the chro- 
matin is the same in amount although now it is in a more irregular nodular 
mass. The milk-white zone is sometimes seen. Some forms apparently 
have two or more nuclei. 161 In the full-grown parasite the chromatin is 
broken up into clusters of fine granules occupying a large achromatic zone. 
Just before segmentation both achromatic zone and chromatin seem en- 
tirely to disappear, the latter later to reappear in fine granules arranged in 
strands and masses throughout the protoplasm, which granules congregate 
into 4 or 5 clusters and then separate into from 15 to 20 dense round masses. 
Achromatic zones now appear around each of these masses. The proto- 
plasm collects around them as a center and the segments separate. 162 
The malarial pigment at the beginning of segmentation is pushed to the 
periphery and, after segmentation is complete, collects in 1 or 2 masses near 
the center. From this it is evident that segmentation is really complete 
before there is any sign of it in the fresh specimen (see page 661). It is 
claimed by many that in the stained specimens the gamete generation 
can be followed from the hyaline form onward . According to Stevens and 
Christophers the young gamete is characterized by the position of the 
chromatin which lies in the center of the vacuole instead of at its edge as is 
the rule in the asexual forms. Some of the infected cells are filled with 

161 For evidence of conjugation, see Ewing, Johns Hopkins Hosp. Bull., 1900. 

162 For much more complex details, see Argutinsky, Arch, f . mikr. Anat. und Entwick- 
lungsges., 1901, Bd. 59, p. 315. 



THE BLOOD 669 

basophile granules which in fresh specimens appear as masses of hemoglo- 
bin suspended in a pale or even colorless stroma. " Plehn's karyochroma- 
tophilic granules," "Schugner's granules" (Plate V, 10, 13). 

Bignami and Bastianelli claim that the division of chromatin into fine 
granules marks the gamete even in the hyaline stage, but Lazear showed 
that this happens always just before segmentation. 

The full-grown macrogamete (Plate V, 14) consists of an abundance of 
deep blue protoplasm and a small compact mass of chromatin peripherally 
placed and surrounded by a thin vacuole-like area. This nucleus occupies 
about }io of the cell. The pigment is uniformly distributed. The remains 
of the red corpuscle often cannot be seen. In the microgametocytes (Plate V, 
11) the chromatin, which is thread-like in nature and in a band arranged 
in a knot or skein, is centrally placed in a large achromatic zone. This 
parasite fills about % of the red cells. Its protoplasm, which is arranged 
in a ring around the nucleus, stains a grayish-green or grayish-red color and 
not the blue of the female form, hence the pigment is more easily seen. 

Quartan (Plate V, 15-18) . — The structure of the quartan resembles that 
of the tertian with the exception that the chromatin mass of the hyaline is 
less dense, is in fact an irregular clump of granules, and that in older stages 
it forms a cluster of fine granules without a distinct achromatic zone, hence 
is often difficult to see. This parasite forms in stained specimens a band 
across the cells. 

^Estivo-Autumnal (Plate V, 19-29). — The chromatin of the sestivo- 
autumnal hyalines is arranged in from 1 to 3 masses or filaments. The 
protoplasm is more scanty than that of the other forms and remains so 
throughout the cycle. Characteristic of this form at a later stage is the 
large oval ring of protoplasm with the thicker layer opposite the chromatin 
mass. The young gamete forms are said to be characteristic (Maurer). 
They are accurately spherical and appear as rings of uniform thickness. 
The nucleus does not project as in the schizonts and the red blood-cell 
usually presents no coarse stippling (Maurer) . The chromatin of the male 
is in a loose network which occupies the most of the cell ; there is compara- 
tively little blue staining protoplasm and the pigment is scattered through- 
out its body. The male crescent is somewhat kidney-shaped and is shorter 
and broader than is the female form. The female crescent is longer and 
narrower, its chromatin more compact and more or less centrally placed. It 
has much more blue staining protoplasm and the pigment is in a ring around 
the nucleus or in a clump near the center. The male circular bodies are 
smaller than the red cells, are perfectly spherical, their chromatin is at first 
in the center in a large irregular mass like a tangled thread and later is in 
4 or 5 dense masses near the periphery. These are extruded as threads, the 
"flagella" (or microgametes) . Sometimes a thin bluish envelop of proto- 
plasm can be stained enveloping this chromatin thread. The macrogamete 
is 2 or 3 times as large as the male form, is often triangular in shape and 



670 CLINICAL DIAGNOSIS 

has abundant blue protoplasm. Its chromatin is in a single mass at the 
periphery and is surrounded by a circle of pigment. 

In the stained specimens (especially in sections cut in paraffin) the 
chromatin mass and part of the protoplasm can be seen to project from 
the surface of the cell, hence the belief (Argutinsky, Stevens and 
Christophers) that the parasite at all stages rests on the cell, not in it (Plate 
V, 20). Study of the fresh blood, their ameboid movements, the way they 
pour through a fine opening when they become extracellular, show, we think, 
that they are intracellular. 

The following points may be emphasized : In the case of tertian and quartan malaria 
the infection must reach a certain intensity (250,000,000 organisms, Ross) before chills 
begin. Because no parasites are found does not rule out malaria, especially if the patient 
has been taking quinine. Fevers with long intervals are explained by the great destruc- 
tion of parasites during each chill, hence several cycles must pass before enough can 
reaccumulate to cause a second chill. 

In a case of fever the discovery of a few crescents in the blood does not mean neces- 
sarily that that particular fever is malaria, since these gamete forms may persist for 
months after the asexual cycle has disappeared. Such patients are malaria carriers. 
But in case hyalines also are found the diagnosis of malaria is justifiable, especially if 
the fever yields promptly to quinine. The asexual cycle is the " febrile cycle." The 
sexual cycle has no direct influence over the host, except that it may again start up the 
asexual. This explains the relapses of malaria in early Spring and especially those which 
follow an accident or a surgical operation even 2 years after any chance of infection has 
passed. Indeed Bass 163 concluded as a result of his survey of the Mississippi Delta that 
between 27.74% an d 37-74% of all persons who have attacks of malaria during a given 
year will relapse during the following year; and that in the case of from 50.77% to 68.86% 
of all persons who have attacks of malaria during a given year it is a relapse and not a 
new infection. 

Not all the members of the same tertian or quartan group are of exactly the same 
size or age and segmentation continues through at least 12 or 14 hours. This is fortunate 
since did they segment more in unison hemoglobinuria would probably be more common 
(as analogue, see Texas fever of cattle). The segmenters vary so in size that it is sup- 
posed that when the majority begin to segment all the others, including those not yet 
quite mature, are drawn into a " precocious segmentation " (Plate III, 16, 17). This 
keeps the groups at an almost equal age, for otherwise these younger forms would dis- 
turb the grouping to the degree which occurs in aestivo-autumnal malaria. This also 
may explain the sudden appearance of a second group in a previously single infection. 
That is, a few of the forms may be so young that they cannot be drawn into segmenta- 
tion and so segment the following day. 

The distribution of parasites in the body is variable. The aestivo-autumnal lives 
for the most part in the spleen, liver and bone-marrow; the same, to a less degree, is 
true of the tertian and still less of the quartan. It is their accumulation in various 
organs which gives rise to symptoms ; if in the brain and medulla they may cause thrombi, 
hence paralyses, transient aphasias, mental symptoms and even sudden death. If in 
the mucosa of the gastro-intestinal tract they cause even necrosis and sloughing, hence 
severe vomiting and diarrhea. 

Whether the severity of an infection depends solely on the number of malarial 
parasites in the body or whether their toxins differ in virulence is hard to answer. It 

163 Contributions to Medical and Biological Research, Dedicated to Sir William 
Osier, in Honor of his Seventieth Birthday, July 12, 1910, by his Pupils and Co-workers. 



THE BLOOD 671 

certainly does not depend on the number in the peripheral blood, although in pernicious 
cases many organisms always are visible. That there is a soluble toxin is proven by the 
degeneration and even wholesale destruction of non-infected cells. This is especially 
true of aestivo-autumnal malaria but also of severe cases of the tertian form. 

TRYPANOSOMIASIS 

Trypanosomiasis, or "sleeping sickness," is due to an actively motile, 
fish-shaped flagellate, Trypanosoma gambiense (Plate II, 21) which can be 
seen in the blood -plasma, moving with a screw-like motion among the red 
blood-cells which it scarcely disturbs. It is from 2 to 3 times as long as a red 
blood-corpuscle (18 to 25^ long, 2 to 2.5^ wide) with 1 flagellum anteriorly 
and an undulating membrane which extends its entire length. 

The parasite should be searched for with a lens of medium power in 
fresh blood specimens, but can more surely be found by aspirating a cervical 
lymph gland or any edematous area. Inoculation experiments may be nec- 
essary. Sometimes many are present in the blood, but generally few. 
They often disappear for long periods, even a month or more, and then 
reappear in force, even 70 to a cover-slip specimen. It may be necessary to 
centrifugalize the blood to find them, in which case the speed of the centri- 
fuge should be slow and the time short, otherwise these protozoa will be 
crushed in the sediment by the red cells. The symptoms of this infection 
seem to bear no relation to the number of parasites in the peripheral blood. 

Stained with a poly chrome-methylene-blue-eosin mixture the protoplasm 
of the body takes a blue stain, the rather large red nucleus is at about their 
middle, a centrosome staining intensely and in a vacuole-like area is very 
near the blunt posterior end and a red line of chromatin runs down the edge 
of the undulating membrane and terminates in the red flagellum. Various 
involution forms will, of course, soon be seen in a fresh specimen. The 
parasite contains ho pigment and hence must live on the plasma. It mul- 
tiplies by longitudinal fission. 

For a long time it was known that similar organisms were a common and 
harmless parasite in the blood of fish, amphibians, birds, rats, and were an 
important cause of disease among horses, cattle and other domesticated 
animals in India, Africa, especially, and South America. The disease bore 
several names. The "tsetse fly disease" of South Africa caused by Try- 
panosoma brucei, which is carried mechanically by a fly, Glossina morsitans, 
which seems to play no part in its life history, is fatal to almost all domestic 
animals, especially the horse, mule, dog, less so for cattle, still less for 
the ass and least for sheep and i goats. Man was, however, supposed 
to be immune. 

The "surra" of India, a disease which attacks horses and camels es- 
pecially, is caused by a parasite discovered in 1881 by Evans, which differs 
in no way from Trypanosoma brucei. The same may perhaps be true of the 
parasite of "mal de Caderas" of Central and South America which attacks 
especially horses. 



672 CLINICAL DIAGNOSIS 

A similar parasite was discovered in 1902 in the blood of man by Dutton 
and in the cerebrospinal fluid of a case of sleeping sickness by Castellani, 
but it was Bruce who first recognized its pathogenic importance in man. It 
was found in cases of sleeping sickness, but also in the blood of some ap- 
parently healthy persons. Of eighty persons in Uganda, all in apparently 
good health, Bruce found it in the blood of 23, but many of these died 
later. The present opinion is that while the human host may be appa- 
rently normal for some time, the disease will sooner or later invade the 
cerebrospinal fluid and cause death. 

This infection can run a rather acute course, but more often it is exceed- 
ingly chronic, causing for years an irregular fever with frequent inter- 
missions, multiple erythema, moderate anemia, marked emaciation, loss 
of strength, localized edema of the face, trunk and legs, enlarged spleen and 
swelling of the lymph glands, especially those of the posterior cervical re- 
gion. Many of the cases resemble malaria. The so-called ' ' sleeping sick- 
ness" begins when the parasite invades the central nervous system. 

To demonstrate this organism in the spinal fluid this should be centrif- 
ugalized gently for fully 5 minutes, and possibly 2 or even 3 times since 
none may be found in the first sediment. The sediment should be examined 
under a well vaselined cover glass. 

The parasite of man, Trypanosoma gambiense, can in no way be dis- 
tinguished either morphologically or pathogenically from that of tsetse 
or surra. 

There are 2 common trypanosomes which are easily distinguished from 
that of man : Trypanosoma theileri, which is pathogenic for cattle alone (a 
parasite from 2 to 3 times as long as the human form), and the trypan osome 
of rats, which is morphologically characteristic since its posterior end is 
long-drawn-out and pointed, and the centrosome is not near the end but at 
the juncture of the posterior and the middle thirds. This can easily be 
distinguished from the other forms even though they coexist in the blood. 
It occurs in about 10 to 30% of rats investigated in some regions, in others 
in even 90%. 

For a good discussion of this whole subject the reader is referred to the 
report by Musgrave and Clegg. 164 

PIROPLASMOSIS 

Leishman-Donovan Disease ; Leishmaniasis. — There are 3 varieties of 
Leishmania : Leishmania donovani,or kala-azar; Leishmania infantum and 
Leishmania furunculosa sive tropica, or oriental sore. 

The Leishman-Donovan Bodies (see Fig. 139), which should be studied 
with the highest magnification possible, are small, oval, round, or cat- 
shaped bodies, from 2.5 to 3.5^ in diameter, with a definite cell outline and 2 

164 Biological Lab. Department of the Interior. Bureau of Government Laboratories 
1903. 



THE BLOOD 673 

chromatin masses, a larger one, the "nucleus," almost round or oval, which 
stains faintly, and a smaller, bacillus-shaped "centrosome," which stains 
deeply and which is directed at right angles, or nearly so, to the axis of the 
nucleus. These 2 bodies are both in the long axis of the cell, the larger 
at the periphery. Many are vacuolated. The outline of the cell cannot al- 
ways be seen, but these 2 masses thus arranged are distinctive. They are 
easily stained by the various polychrome methylene blue-eosin mixtures. 

They have been found in the circulating -blood only of fatal cases, but 
are easily demonstrated in blood obtained by splenic puncture and also 
in the granulation tissue snipped off with scissors from the ulcers of the dis- 
ease and crushed thin in the slide. At autopsy many are found in the 
mesenteric lymph-glands, bone-marrow and liver. 

Grown in proper media (e. g., McNeal-Novy-Nicolle agar) these develop 
into club-shaped flagellates which have no undulating membrane. 

Some lie free, but some seem to be intracellular; one or two in a leuco- 
cyte ( ?) , from 1 to 1 2 in endothelial or splenic cells and even hundreds in 
macrophages (?). These last masses are variously interpreted. If cells, 
they are badly degenerated. Ross considers them to be a "matrix" in 
which the organisms lie and thinks that none are intracellular. Manson 
regards such masses as zooglia. 

Their division begins in the larger chromatin mass and ends in the 
smaller which may begin to divide after the fragments of the larger are 
widely separated. 

This parasite is supposed to be the cause of some cases of chronic "ma- 
larial" cachexia of the Tropics, of dum-dum fever, kala-azar, tropical 
splenomegaly of the tropical ulcer, Delhi boil, Aleppo button, Scinde sore, 
oriental sore, etc. It is a filth disease. In the Tropics it promises to prove 
even more important than the malarial organism. 

Donovan 165 reports 7 2 cases with a mortality of 30. 5 5 %. Clinically one 
finds great enlargement of the spleen, emaciation, irregular fever, various 
abdominal symptoms, cutaneous hemorrhages and ulcerations. 

The blood features are a moderate anemia, from 2,000,000 to 4,000,000 
red blood-cells, and a leucopenia with a relative and absolute increase of the 
large monconuclears. The average leucocyte count is about 2,000. In a 
case of Neave, an 8-year-old boy with a total count of 3000, the l.m. were 
67%, pmn.n. 20%, s.m. 11%, eos. 1% and myelocytes, 1%. In most cases, 
however, the formula is more nearly that of normal blood . 

FILARIASIS 

Of the various forms of filaria, the embryos of which are found in human 
blood, the most common is Filaria bancrofti (F. nocturna). The embryos 
of this filaria are from 270 to 340M long, and from 7 to ii/j. broad (see 
Figs. 140 and 141). They are enclosed in a sheath which is considerably 

165 Lancet, September 10, 1904. 
43 



674 CLINICAL DIAGNOSIS 

longer than the parasite. The anterior end of the worm is abruptly- 
rounded, with a six-tipped prepuce and a sharp fang; the posterior tapers 
off for % of its length. It has a granular median axis. 

Their movement at first is progressive, but their anterior end seems soon 
to become attached to the glass and there they remain for days lashing the 
surrounding corpuscles. These embryos appear in the circulation towards 
evening. Their number gradually rises to a maximum at about midnight 
then diminishes towards dawn. During the day they are in the internal 
organs, especially the lungs. An estimated number of 40 or 50 millions of 
microfilaria circulating in the blood may give rise to no symptoms since 
their size permits them to traverse the capillaries unobstructed. These 
embryos apparently have no pathogenic properties whatever, but the parent 
worm and the immature products of conception are dangerous. 

Lothrop and Pratt 166 made an hourly chart of the number of embryos 
present. In that case the maximum was at midnight when there were 2 100 
per c.cm. of blood. 

The adult worms lie in the thoracic duct or smaller lymphatic trunks obstructing 
the lymph flow and causing lymphedema of the undrained area, therefore lymph-scrotum, 
varicose groin glands and various other lymph tumors. If a lymphatic varix develops 
near the kidney, bladder, testicle or peritoneum and ruptures, chyluria, chylocele or 
chylous ascites may result. This obstruction may in part be due also to the eggs, which 
measure from 28 to 30jii long and 15^ wide, therefore are too large to pass through lymph 
capillaries. They explain most cases of hematochyluria and also of elephantiasis, in 
which condition one seldom finds microfilariae in the blood. In these cases it is supposed 
that the female parent worm located in a lymphatic vessel or lymph gland of the affected 
part has aborted her immature ova which, since they have a larger diameter than the 
healthy fully formed microfilarias, cause embolism of the lymphatic glands resulting in 
great enlargement or elephantiasis of the leg, arm, breast, scrotum or vulva. The 
female is from 85 to 150 mm. long with a distinct neck, a head with simple minute 
terminal mouth, a plain cylindrical body which tapers toward the neck, and tail which 
is covered by a striated cuticle. The tail ends bluntly and has a small depression sur- 
rounded by 2 lips. The anus is a ventral opening on the summit of a trilobed papilla. 
The females are generally viviparous, each giving birth to about 1000 embryos which 
live probably for years in the blood. The embryos reach the general circulation through 
the thoracic duct. 

The male is 80 mm. long, has no neck but has a tendril-like tail rolled up in 1 or 2 
spirals. The esophagus is thick walled. The cloaca is ventral, with 4 pre-anal and 
4 post-anal papillae and 2 spicules. 

The intermediate hosts of filaria are some varieties of mosquitoes. Some of the 
known efficient hosts of Filaria bancrofti are Culex fatigans, Stegomyia pseudoscuttelaris 
and Anopheles maculipennis. About an hour after the bite the embryos in the mosquito's 
stomach cast their sheaths. Some die, but others bore their way actively through the 
intestinal wall to the muscles, where they rest. During the next 2 or 3 days the embryo 
becomes larger and its alimentary tract develops. There is no increase in their number 
in the mosquito. On the seventh day the worm is 1.5 mm. long and perfectly developed. 
It now actively travels to the head and takes its position in the labium (Fig. 142), 
whence it enters its new host during the biting process by piercing the delicate membrane 

166 Am. Jour. Med. Sci., November, 1900. 



THE BLOOD 675 

of the end of the proboscis, dropping on the surface of the skin and boring its own way 
through to a subcutaneous lymph channel. It requires an infection by even hundreds 
of these adult forms to cause a very severe case and it may be years before any symptom 
begins. At least I male and female must locate in the same lymph gland and develop 
simultaneously to produce embryos. 

The clinical symptoms, in addition to the various lymph tumors, are anemia, 
enlarged spleen and fever. In any case of lymph tumor, elephantiasis, hematochyluria, 
the blood should be examined. These cases are usually admitted to the surgical side, 
and some have been operated for inguinal hernia, the lymph-scrotum being thus 
interpreted. One of the chief foci in this country is Charleston, S. C. Probably there 
are a good many cases in this country, judging from the number found in quite widely 
distant cities. 

Francis 167 illustrated the difficulty with which this disease is spread as follows : 
Nine cases of filariasis found in cities outside of Charleston showed an estimated average 
of only 124 microfilariae per cubic centimeter of blood. Supposing that an infected 
individual harbors 124 microfilariae per cubic centimeter of his blood and that a mosquito 
draws 1 . 1 5 mgms. of blood at a bite, a mosquito would then have to bite such an infected 
individual 7 times in order to get 1 microfilaria ; or, if 7 mosquitoes bit him, only 1 of the 

7 would imbibe a microfilaria. If this 1 microfilaria encountered no obstacles whatever 
in its cycle, either in the mosquito's stomach, in the thoracic muscles of the mosquito 
or in its proboscis, or on the human skin or in the lymphatics of the man, it would 
successfully reach a lymph-gland. Here its activities might, however, come to an end 
by reason of the absence of 1 of the opposite sex in that same gland. 

The second set of 7 mosquito bites, if successful, might result in the deposit of a 
second filaria in some lymph-gland of the body but quite distant from the first one. 
It would not be difficult to distribute 10 infective mosquito bites over the body 
surface in such locations that no 2 of the 10 implanted filariae would meet in the 
same gland. 

If a male and female did lodge in the same gland and the female gave birth to the 
average 1000 embryos, then since there would be no further supply of embryos, 5217 
mosquitoes would have to fill themselves of this patient's blood to get 1 single embryo. 
That is, for the spread of this disease, mass blood infection 'and mass mosquito biting 
both are necessary. 

Filariasis occurs endemic in the Tropics where it is called "craw-craw," 
or the "sleeping disease." In the Fiji Islands as many as 25% and in the 
Friendly Islands even 32% of the inhabitants are said to carry this infection. 

The blood should be examined late at night. A very thick fresh 
specimen is made and examined with the low power. These worms 
cannot be overlooked. Their motion will continue for even a week in a 
well-sealed specimen. 

Francis recommends to obtain the blood between 10.30 p.m. and 1.30 
a.m. by sticking, with a Hagedorn needle, the tip of the finger. From 6 to 

8 large drops of blood are squeezed out and allowed to fall on an ordinary 
clean glass slide. With a fresh toothpick the blood is evenly spread over 
the most of the surface of the slide before it has had time to clot. The 
thick blood films are dried in the air in a level position, protected from flies 
but freely exposed to the air. As soon as they are dry the slides are stood 

167 Hygienic Lab. Bull. No. 117, June, 1919. 



676 CLINICAL DIAGNOSIS 

upright, back to back, in Coplin jars containing preferably distilled water, 
or good tap water. To get a colorless film it should be dehemoglobinized 
as soon as dry. An hour usually suffices for the water to entirely remove the 
hemoglobin, leaving the film colorless but otherwise unaltered. If desirable 
the slides can be transferred to fresh water to complete the laking. The 
slides are examined while still wet on a mechanical stage. In such prepara- 
tions the glistening microfilariae are readily seen. In routine work where a 
determination of the number of individuals infected is the sole object of the 
investigation, examination of the wet laked films is entirely satisfactory 
and was followed in the government surveys. 

If the specimens remain wet 48 hours while awaiting their turn for 
examination, bacterial growth is likely to disintegrate the film and cause its 
detachment from the slide. A few drops of formalin added to the water 
will prevent this. 

To stain the microfilariae thick smears decolorized in distilled water as described 
above are made. They are sufficiently fixed to the slide dried in the air. They are then 
stained with Delafield's hemotoxylin for 5 minutes, steaming them over a flame, as in 
staining tubercle bacilli, and washed thoroughly in tap water They are next decolored 
with acid alcohol sufficiently to differentiate with the low power the cells of the cell 
column of the parasite, dehydrated in absolute alcohol, cleared in oil of cloves or xylol 
and mounted in Canada balsam. 

The polychrome stains commonly employed in blood examinations 
may also be used. 

Hematochyluria is due to rupture of the varicosed lymph- vessels of the 
bladder, these forming part of the collateral circulation which compensates 
for an occluded thoracic duct. Attacks of this may recur for even 18 years, 
each being weeks or months long and separated by intervals of months or 
years during which the urine is clear. The attacks come on with pain and 
fever, spontaneously or following exertion, excitement, etc. The sequence is 
hematuria, hematochyluria and chyluria. The urine contains most blood 
and most embryos in the early morning but most chyle after a rich meal 
(even 3.8% fat). 168 

Filaria Diurna (the embryos) differs little from Filaria nocturna (bancrof ti) , except 
that they remain in the circulation only during the day. The granular axis is lacking 
The adult form is not yet known (Filaria loa of the subcutaneous areolar tissue?). 

Filaria Perstans. — These embryos which remain in the circulation day and night 
are about 200yu long and 4 to 5/z in width, have no sheath, a body which tapers for its 
posterior % and a slightly bulbous tail. They make a very active, progressive, as well 
as a lashing motion. The adult is found in the retroperitoneal tissue. 

Other forms described in man are Filaria ozzardi (embryos small, 170 to 200/x long, 
without sheath and with sharp tail, no periodicity and its adult in the sub-peritoneal 
tissue) ; Filaria demarquai (embryos 200fd long, sheathless and sharp-tailed, with cephalic 
armature, no periodicity and the adult doubtful); Filaria megalhesi, Filaria gigas and 
Filaria loa. 

168 p or the blood formula, see page 561. 



THE BLOOD 677 

RELAPSING FEVER 

The parasite of relapsing or famine fever of Europe is Spirochete (ro 
Spirillum) obermeieri, an organism curled like a corkscrew, from 12 to 45m 
long and from 0.3 to 0.5/j in breadth (Fig 143). Its curves, from 4 to 16 in 
number, are sharp and regular. Its ends are pointed. It is flagellated, but 
there is a dispute as to the number of flagella. This organism is present in 
the circulating blood from the onset of the fever until the crisis when it 
suddenly disappears. At the beginning of the attack the parasite moves 
with a rapid corkscrew motion among the blood corpuscles, which are not 
much disturbed, but later there is simply an undulatory movement of the 
whole spirochete and still later merely a slight swaying motion. After the 
crisis, it is said, the parasites collect in the spleen. Little is known of the 
life history of this organism outside the human body. Relapsing fever is 
certainly a filth disease and the bed-bug is accused of being the agent of 
its natural transmission, but this is not yet proved. 

Three similar organisms have been discovered. 169 These are Spirillum 
duttoni, the cause of the African disease; Spirillum carteri, cause of the 
Asiatic disease; and Spirillum novyi, cause of the American disease. 

Spirillum duttoni varies in length from 15 to 45/x and from 0.2 to 
0.41J, in width. It has from 2 to 6 curves. Whether it has flagella and un- 
dulating membrane is disputed. Not nearly as many of these spirilla are 
present in the blood during these attacks as in the case of the other spirillar 
fevers. Transmission to man seems to be by ticks. 

Spirillum novyi is an organism whose length is from 7 to q/jl or mul- 
tiples of this. It is shorter and finer than the other spirilla and has 2 or 
3 sharp regular curves. 

Spirilla carteri is a parasite from 12 to 16 /jl long and from 0.3 to 
0.5/x wide. This organism causes a severe infection. Lice are supposed to 
spread this disease. 

Trichinella spiralis was found in the circulating blood first by Herrick 
and Janeway, 170 who used Staubli's method. The blood, from a few drops 
to 10 c.c. in amount, is laked with from 10 to 15 parts of 3% acetic acid. 
This laked blood is centrifugalized and the sediment examined. 

169 Mackie, N. Y. Med. Jour., Aug. 2, 1908. 

170 Arch, of Int. Med., April, 1909. 



CHAPTER VI 

CEREBROSPINAL FLUID 

The normal amount of cerebrospinal fluid in the subarachnoid space and 
in the ventricles is said to vary from 60 to 150 c.c, of which from 5 to 10 c.c. 
or more may be withdrawn by puncture without untoward symptoms. 
The technic of lumbar puncture cannot be described here further than to 
say that a monometer, if only a long glass tube with barometer bore, should 
be attached to the needle, and the pressure frequently noted during each 
puncture. This pressure normally varies, the patient recumbent, from 120 
to 1 80 mm. of water in the adult and from 45 to 90 in the child. The amount 
to be removed will depend partly on the drop in pressure caused by the loss 
of the first 5 c.c. of fluid removed, but more on the provisional diagnosis. 
In any doubtful case it is a safe rule never to remove over 5 c.c. at a time, 
for in patients with brain tumor sudden death has sometimes followed 
lumbar puncture. 

It is relatively most abundant in the first years of life. In cases with 
traumatic fistula of the spinal canal a flow of from 1^2 to 2 liters in 24 hours 
has been observed and in cases of cerebrospinal rhinorrhea the amount which 
has flowed from the nose has varied from 96 to 720 c.c. per day. These 
figures give some idea of the rapidity with which this fluid can be secreted. 
It is pathologically increased in amount in certain infectious diseases, in 
meningitis, always in hydrocephalus and in general paralysis of the insane. 
Coriat was able to obtain by puncture in alcoholic cases from 10 to 100 c.c, 
in dementia precox even 50 c.c, and in general paralysis sometimes over 
100 c.c. In senile cases one gets even 60 c.c 

Levinson recommends that the fluid be collected in several test-tubes of uniform 
width, the amount removed to depend on the amount of fluid present as indicated by 
the pressure under which it flows. 

The first few drops are allowed to escape to make sure the fluid is free of blood. Two 
cubic centimeters are collected in the first test-tube and is used for (1) the cytologic 
examination, (2) a direct smear for microorganisms, (3) cultures, (4) 1 c.c. for the per- 
manganate test. These tests should be made at once. 

From 3 to 5 c.c. are collected in the second tube. In case the pressure is not increased 
the contents of this tube should be used for the globulin tests, the Wassermann and the 
Lange tests. If the pressure is increased this tube is set aside for the formation 
of a pellicle. 

If there is enough fluid for the third tube, from 5 to 8 c.c. are collected for (1) the 
second of the permanganate tests, (2) the Ross- Jones, Noguchi, Nonne and the sulpho- 
salicylic-mercuric chloride tests, (3) Lange's test, (4) the Wassermann test, (5) the sugar 
content, and (6) centrifugalization for the examination for organisms. 

Into the fourth tube is run any fluid to be collected over that in these three tubes. 
678 



CEREBROSPINAL FLUID 679 

In color the fluid is either absolutely limpid or has a slight yellowish 
color due to lutein, the pigment of blood-serum. Xanthochromia, or a 
yellowish color of the spinal fluid, is present in many cases of tumor of the 
cord, especially those of the cauda equina and conus medullaris, often in 
extradural compression of the cord, in old cases of brain hemorrhage while 
in some cases it is due to the trauma of a vein received at a previous lumbar 
puncture. The cerebrospinal fluid of patients who have had a subdural 
hemorrhage may be red, of jaundiced patients a greenish-yellow, while 
the presence of pus will give it an opaque yellow color. It may also be 
stained by certain drugs, as, for instance, methylene blue. 

No pellicle or sediment will appear in a normal fluid properly protected 
from contamination. 

A cloudy fluid generally means pus, while fluids from cases of tuber- 
culous meningitis are beautifully clear at first but soon show a fine spider- 
web clot of fibrin strands. If a clear fluid is not kept sterile it will quickly 
become turbid from bacteria. 

In reaction this fluid normally is alkaline, but even in as short a time as 
10 minutes after death it may rapidly become acid. Some have claimed 
that it may be acid during life because of the lactic acid formed by the 
fermentation of decomposing cells. In i case lactic acid (inactive) is said to 
to have been present after epileptiform convulsions. Its reaction depends 
directly upon the reaction of the brain tissue. 

The specific gravity of the cerebrospinal fluid, Coriat states, is normally 
from 1.007 to 1.010 (Halliburton, 1.006 to 1.008). While this varies much in 
different diseases nothing specific has as yet been determined. In general 
paralysis 1.009 "to 1.012 are common figures; in hydrocephalus 1.008 to 
1.009. These last figures are within the normal limits. In a case of 
spinal bifida it was reported to be 1.001. 

In the cerebrospinal fluid is found from 0.04 to 0.05% of a reducing 
body the nature of which was for a long time in dispute. It was reported 
to be galastose, pyrocatechin or a xanthin body but lately it has been iden- 
tified as dextrose. It is increased in amount by repeated tappings. 

Determination of Glucose in the Spinal Fluid (Lewis and Benedict's 
Method). One measures 2 c.c. of the cerebrospinal fluid into a 25 c.c. volu- 
metric flask containing 5 c.c. of distilled water. To this are added 15 c.c. 
of saturated picric acid solution and water up to the 25 c.c. mark. This 
mixture is then well shaken and filtered. One measures 8 c.c. of this filtrate 
into each of 2 (the 1 for a control) large test tubes of Jena glass, adds to 
each 2 c.c. of saturated picric acid solution and 1 c.c. of 10% sodium carbo- 
nate solution and evaporates the contents over the free flame until 
precipitation occurs. Three cubic centimeters of water are then added and 
the tubes heated again to the boiling point to dissolve the precipitate. The 
contents of the tubes are then transferred to two 10 c.c. volumetric flasks, 
cooled, the flasks filled to the 10 c.c. mark with water, shaken and filtered 



680 CLINICAL DIAGNOSIS 

through cotton into a colorimetric flask and the color compared in a Du- 
bosque colorimeter with either a glucose standard made up fresh each 
time or with a permanent standard solution containing 0.064 nig. of 
picramic acid and o. 1 gm. of sodium carbonate in 1000 c.c. of water. 

Mott and Halliburton found this body absent in 12 of 14 cases of general 
paresis. It is usually absent in tuberculosis and in epidemic cerebrospinal 
meningitis. Others claim that glucose is increased in hydrocephalus (Cav- 
azzini), diabetes (Schaefer, in which case from 0.32 to 0.35% is said to 
have been found) and in grave pneumonia. 

Normal cerebrospinal fluids have been found to have the fol- 
lowing composition. 1 

In 1000 parts they were: 

Fixed matter 10.65 -11.00 Average 10.93 

Organic matter 1.75 - 2.65 " 2.13 

Mineral matter 8.50 - 9. " 8.80 

Protein 0.13 - 0.30 " 0.18 

Amido acids " 0.010 

Urea 0.03 - 0.1 " 0.06 

Total N 0.196- 0.198 " 0.197 

[Non-proteid N 0.17 - 0.26 " 0.21] 

[Creatinin 0.007- 0.015 " 0.009] 

Reducing bodies 0.48 - 0.58 " 0.53 

[Sugar 0.7 - 1. " 0.7] 

Organic acids " 0.30 

Chlorides 7.25 - 7.40 " 7.32 

Total phosphates (as P2O2) 0.029- 0.031 " 0.030 

Inorganic phosphates (as P2O5) 0.012 

Organic phosphates (as P2O5) 0.018 

Total sulphur (as SO) 0.028- 0.071 " 0.056 

[Inorganic sulphur (as SO) 0.010] 

[Organic sulphur (as SO) 0.046] 

Nitrates 0.009 

Sodium as Na 2 4-346 

Potassium as K2O 0.251 

Calcium as CaO 0.095 

Magnesium as MgO 0.050 

The bracketed figures are from other calculations. 

Le vinson, who determined the H-ion concentration in over 400 normal 
fluids found that immediately after the fluid was withdrawn from the body 
Ph ranged from 7.4 to 7.6 or exactly the same as that of the fresh blood. 
After standing 30 minutes Ph became 7.5 to 7.6, and in 12 hours 8.1. 

He found the alkali reserve (van Slyke method) in non-meningitic 
fluids varied between 45.7 and 63. (or about the same as that of the blood. 

The urea may be increased in hydrocephalus; much, even to 0.45% 
in nephritis and considerably in arteriosclerosis. 2 

1 Levinson Cerebrospinal Fluid in Health and in Disease. 19 19. 

2 Widal and Froin, Gaz. des Hop., No. 122, 1904. 



CEREBROSPINAL FLUID 681 

The chief proteid found in normal fluid is globulin. Quincke estimates 
the total proteid as from 0.2 to 0.5, Ricker from 0.5 to 1, and Gumprecht 
0.25 gm. per liter. Halliburton found globulin alone in 13 cases and in 6 
cases albumose also, while in 3, 2 of which clearly were cases of meningitis, 
albumin was also found. No fibrinogen is ever found normally. If a little 
blood-serum is added to the cerebrospinal fluid its presence will be evidenced 
by fibrin formation. Serum albumin is said never to be present normally. 
In meningocele albumose and peptone have been found. 

In general paresis the total solids may reach even 2.39 p.M. and the 
proteids are considerably increased. There is some increase in hydroceph- 
alus, in inflammatory conditions, in cases with stasis due to brain tumor 
(2 to 4 p.M.), after repeated tappings, in apoplexy and in meningitis (as a 
rule, 2 to 3 p.M. ; but if purulent 7 to 9 p.M.). 

Mott and Halliburton recommend the following quantitative method 
for total protein. The fluid is made acid with acetic acid and 2 volumes 
of absolute alcohol are added. It is then boiled, filtered, the precipitate 
dried at 1 io° C. and weighed. 

In 8 cases of general paresis the average percentage was 0.239% and in 2 
of spina bifida, 0.088%. In general paresis the proteoses and peptones 
were absent, but globulin and a little nucleoproteid were found. 

To demonstrate nucleoproteid in the spinal fluid it is necessary to use at 
least 1 liter of the fluid. To this an excess of alcohol is added and the pre- 
cipitate digested in water. If the undissolved residue is found to contain 
a high percentage of phosphorus, suggesting nuclein, the residue is washed 
with 0.2% HC1 and heated on the water-bath at ioo° C. with fuming 
HNO3 and a small amount of H 2 S0 4 and KCIO3. The residue is dissolved 
in HNO3 and ammonium molybdate added; a yellowish crystalline precipi- 
tate will result. 

A very good idea of the amount of protein present may be gained by the 
heat-acetic-acid test. A normal fluid would remain clear on standing some 
hours. If a fluid in which the protein is increased be boiled no cloud will 
appear, but on adding one drop of dilute acid a very faint white opales- 
cence develops, which will separate in fine flocculi. If some of this fresh 
fluid is mixed with an equal amount of saturated ammonium sulphate so- 
lution the globulin will be precipitated. This is removed by filtration, 
the clear filtrate is then acidified with acetic acid and boiled. The pre- 
cipitate will be albumin. 

Globulin Tests.— These tests are of value only if the spinal fluid is 
free of blood. 

a. Noguchi's Test. — One measures into a small test tube with a pi- 
pette 0.2 c.c. of the cerebrospinal fluid, adds 0.5 c.c. of butyric acid solution 
(5 c.c. of butyric acid in 45 c.c. of physiologic salt solution), boils this mix- 
ture for a few seconds, then adds o. 1 c.c. of a 4% aqueous NaOH solution 
and boils again for a few seconds. A definitely flocculent precipitate, 



682 CLINICAL DIAGNOSIS 

whether of fine or coarse flocculi, which appears in from 5 to 20 minutes, is 
proof of an increase of globulin in the fluid. A normal fluid would remain 
clear for at least 2 hours or become only slightly opalescent. This is the 
most accurate of the globulin tests. 

b. The Ross-Jones Test. — One superimposes 0.3 c.c. of the spinal 
fluid on an equal amount of saturated ammonium sulphate solution. The 
appearance of an opaque ring at the point of contact of the 2 fluids indicates 
an increase of globulin. 

c. The Nonne-Apelt Test. — One mixes equal amounts of cerebrospinal 
fluid and saturated ammonium sulphate solution. A white precipitate 
appearing in 3 minutes is positive for euglobulin. One now filters out the 
precipitate, adds one drop of 10% acetic acid to the filtrate and boils this 
fluid. A precipitate is evidence of serum albumin. 

d. Kaplan's Method. — One heats to the boiling point (twice) 0.5 c.c. 
of the spinal fluid and then adds 3 drops of a 5% solution of butyric acid in 
physiological salt solution and immediately 0.5 c.c. of a supersaturated 
solution of ammonium sulphate. This mixture is now set aside for 20 
minutes. An excess of globulin is indicated by a thick granular precipitate. 

e. Pandy's Method. — One drop of cerebrospinal fluid is added to 1 c.c. 
of a concentrated solution of carbolic acid (1 part of phenol crystals to 
15 parts of water). A bluish-white ring or cloud will indicate an ex- 
cess of globulin. 

One objection to this test is that it is so sensitive that it may appear 
positive in the case of normal fluids. 

f. The Sulphosalicylic-Mercuric Chloride Method. — One meas- 
ures 1 c.c. of cerebrospinal fluid into each of 2 similar small test-tubes about 
0.3 cm. in diameter. To one is added 1 c.c. of 3% sulphosalicylic acid, to 
the other 1 c.c. of 1% mercuric chloride solution. Both tubes are then set 
aside for 24 hours, at the end of which time the precipitates in the 2 tubes 
are compared. In the case of normal fluids the sediments in both are very 
slight. In all cases of suppurative meningitis, however, the precipitate in 
the tube with sulphosalicylic acid is even 3 times as abundant as that in the 
other, while in tuberculous meningitis the reverse is true. This last point 
has been found very valuable. 

The Permanganate Test. — This test, introduced by Mayerhofer, for 
the total organic content of cerebrospinal fluids is a modification of the 
standard test used for a similar purpose in the analysis of drinking waters. 

One measures with an accurate pipette 1.0 c.c. of the cerebrospinal fluid 
into an Erlenmeyer flask and adds 50 c.c. of distilled water and 10 c.c. of 
dilute H 2 S0 4 (1 part H 3 S0 4 in 3 parts of water) . This mixture is then 
brought to the boiling point, to c.c. of 0.1 N KMn0 4 solution added and 
the boiling continued for exactly 10 minutes. Then 10 c.c. of 0.1 N 
oxalic acid are added and the 0.1 N KMn0 4 run in drop by drop from a 
burette until the red color persists throughout the body of the fluid for 



CEREBROSPINAL FLUID 683 

several minutes. In this way one determines the amount of the 0.1N 
KMn0 4 used up in oxidizing the organic matter. 

One next determines the amount of o.i N KMn0 4 necessary to oxi- 
dize the organic matter in the reagents used by repeating the above pro- 
cedure, only without adding any cerebrospinal fluid. The result is to be sub- 
tracted from the first reading. The difference is the permanganate index. 

The saline constituents of cerebrospinal fluids resemble those of other 
serous fluids. 

The toxicity of the fluid has been found increased in general paresis and 
also after epileptic seizures. Its poisonous qualities are due to cholin 
and other products of nerve degeneration. 

Cholin is a decomposition product of lecithin, the chief component of 
the myelin sheaths. Its appearance in the spinal fluid is evidence of nerve 
disintegration. It is soluble in water and alcohol, insoluble in ether and is 
precipitated by PtCl 6 as polymorphous crystals, which, however, if re- 
crystallized from warmed 15% alcohol are regular octahedra. These crys- 
tals are insoluble in alcohol and ether, but are soluble in water. 

The careful technic given by Coriat for the demonstration of cholin in the spina 
fluid is as follows: The proteids are first precipitated by 95% alcohol in excess and the 
filtrate evaporated over the water-bath at 40 C. to dryness. The residue is extracted 
with absolute alcohol, filtered again and this evaporated to dryness. This process is 
repeated several times, the temperature always being kept low. All traces of proteid 
and potassium salts are thus removed. The final residue, after extraction with absolute 
alcohol, is a syrup of a light color. This is divided into 2 fractions. The first is dissolved 
in distilled water, and the second in 15% alcohol. The watery solution is tested for 
proteid by the biuret, Millon's and other proteid reactions all of which must be nega- 
tive. It is tested also for cholin by the ordinary reactions for alkaloids (phosphotungstic 
and phosphomolybdic acids, et. al.) all of which must be positive. To the alcoholic 
solution are then added 4 drops of 4% PtC1 6 . It is then evaporated in a watch-glass 
over CaCl 2 to obtain the crystals. The presence of cholin may be assumed if among 
the former tests tannic acid gave no precipitate (thus neurin is excluded) and precipi- 
tates were obtained with phosphotungstic acid (white) , phosphomolybdic acid (yellow) , 
PtCl 6 , AuCle, and with Lugol's (brown). On evaporating the 15% alcohol solution large 
yellow octahedral crystals must separate which are easily soluble in water (therefore 
they are not neurin). Their size, solubility in water and the fact that the aqueous 
solution gave the alkaloid reaction, excludes potassium. These crystals, if in a sufficient 
amount, may be dried and the platinum in them determined, which should be 34.8%. 

In the same hydrolysis of lecithin are formed glycerophosphoric acid 
and stearic acid. The latter unite with the glycerol radicals to form the 
neutral fats upon which Marchi's stain depends. 

Cholin is eliminated in the cerebrospinal fluid and the blood and 
glycerophosphoric acid in the urine. 

The presence of cholin indicates nerve disintegration. It has been found 
in a wide variety of nervous disturbances: in general paresis, combined 
sclerosis, insular sclerosis, alcoholic neuritis, beriberi, senile dementia, 
delirium tremens, etc., and in amounts roughly parallel to that of proteids 



684 



CLINICAL DIAGNOSIS 



present. Mott considers that its presence or absence can be used to 
differentiate the organic from the functional nervous disturbances only 
in case the organic disturbance was active at the time the fluid was 
obtained. It is most constantly present in general paresis, in which 
disease Coriat found it in all of 14 cases. He found, however, no relation 
between its amount and the anatomical rinding. 

We add a table of a few analyses we had formerly made, calling particular attention 
to the high solid content in stasis (due to brain tumor), and to the difference between 
the ventricular and spinal fluids in a case of hydrocephalus, a difference whicn we had 
noted in 2 previous cases. 

Cerebrospinal and Ventricular Fluids 






6 


Case 




- '6 
< 


> 

u 
bo 

ft 
W 


c 



u 

ft 

(0 

'o 
w 


a 



!-. 
ft 
[0 

rti 

'0 


i-. 
ft, 


Salts and Extrac- 
tives soluble in 
H2O; insoluble 
in alcohol. 





ft 

a 
13 

H 


Extractives and 
salts soluble in 
alcohol; urea, 
sugar, NaCl, etc. 


c 




u 

ft 

,4 

a 


15 +j 

• -S c 
+j <-> 

c8 - O 

- 1- O 

.Hi! a 

-'Cm - 
woo,; 
P.07; H 

o3 >.j2 

woo 


I 


Normal child 

Normal child 

Hydrocephalus cord . . 


13 

22 


IOO7.4 
IOO8.3 
1002. 
I006.2 
















2 
















3 
















Brain (fluid from) . . 




0.96 


O.O99I 

O.I70 

0.III2 


O.2703 

O.62 

O.5964 


O.507 

O.492 
O.5166 


O.2937 
0.374 






4 


Hernia of brain (tumor) 
Later 


50 
630 
450 

200 

25 
100 

25 

10 

30 


0.2092 


0.023 

0.016 


IOO6.9 
IOO7.7 

1011.6 

1009.2 

1007. 

1009.2 
1018.8 

1006. 
1008. 


2.5132 




Later 








5 
6 


(Ventricular fluids) 
Tumor of brain ...... 

(Hernia) 
Gunshot wound; head 
Cerebrospinal menin- 
gitis 


















2.664 


O.7839 


O.68 


O.I988 






7 






8 


Streptococcus menin- 
gitis 




O.I628 
O.066 












9 
10 


Tuberculousmeningitis 
Pneumonia ; meningeal 

symptoms 

Paresis? 


O.5629 


O.4712 


O.4H2 


0.2445 


0.0185 


1 1 




0.059/ 























Cytology. — To count the cells of a spinal fluid the tube of a leucocyte 
pipette is filled first to the 0.5 mark with some staining fluid, e. g., Unna's 
polychrome methylene-blue, and then to the eleven mark with the perfectly 
fresh spinal fluid. This is well shaken and allowed to stand a few minutes 
that the cells may become stained. The shaking must be repeated immedi- 
ately before the count is made. While an ordinary leucocyte counting 
chamber may be used, more accurate results are obtained with the special 
Fuchs-Rosenthal chamber which is 0.2 mm. deep and the sides of the ruled 
square of which are 4 mm. long. The volume of fluid bounded by this 
square will be 3.2 c.mm. If A = the average number of cells in the squares 
10a 



counted, then 



3 A 



number of cells in 1 c.mm. of the diluted fluid. A 



CEREBROSPINAL FLUID 685 

correction of 5% may be made for the slight dilution. The average number 
of cells in normal fluids lies between 4 and 6 cells per 1 c.rnm. Above 6 is 
suspicious, while above 10 indicates a distinctly pathological condition. 

The cells in normal fluids are small mononuclear leucocytes similar to 
those of the blood, together with, very rarely, a few large mononuclears. 

In case the fluid cannot be examined while fresh, it may later be centrif- 
ugalized for 1 5 minutes and the sediment obtained spread on a slide in a 
round smear about 7 mm. in diameter, which is then fixed and stained. 
In the case of normal fluid not over 4 lymphocytes will be found in each 
field of a 400 magnification. 

The most interesting application of the cell count of spinal fluids is in 
the diagnosis of cerebral lues, general paresis, and tabes dorsalis, diseases 
with a chronic posterior meningitis and a slight, though usually definite, 
spinal leucocytosis. In general paresis a lymphocytosis and an increase of 
the protein of the spinal fluid may be the earliest symptoms. A negative 
find is often of more value than a positive one. In 80 cases the counts varied 
from 5 to 204 cells per 1 c.rnm. In 9 of these the count was under 5, in 
6 over 100 (Rous). In 25 cases of this disease Cornell 2 found the count to 
vary from 12 to 216 cells, the average being 52. Of these cells, from 45 to 
97% were small lymphocytes, from o to 15% (average 4%,) large lympho- 
cytes, from 1 to 56% (average 18%) polymorphonuclear neutrophils and 
from 0.1 to 5% (average 1.5%) plasma cells. In the diagnosis of cerebral 
lues the cell counts have thus far been of little aid. A slight spinal lympho- 
cytosis has been found present in about half the cases of locomotor ataxia 
studied and the same is true of some cases of multiple sclerosis. On the 
other hand, in a large group of mental and nervous diseases, among which 
are included the psychoses of arteriosclerosis, chronic alcoholism, chronic 
delusional states, dementia precox, epilepsy, most manacal and hypo- 
maniacal conditions, the psychoneuroses, uremia, hydrocephalus and cere- 
bral neoplasm, the cytology of the fluids have thus far proved negative. 

Bacteriology of the Spinal Fluid. — To determine the organism present, 
smears of fresh fluid should be stained and cultures made, preferably on 
blood agar. Among the organisms most often found in cloudy fluids are: 
Micrococcus intracellularis meningitidis, Diplococcus pneumoniae, Bacillus 
influenzae, Streptococcus pyogenes, and several other organisms (e. g. f 
Bacillus typhosus, Bacillus coli, Bacillus paratyphosus, Bacillus mallei 
Bacillus pestis, the Gonococcus, staphylococci, etc.). In the clear fluid 
of tuberculous meningitis Koch's organism may be demonstrated. 

Lang's Gold Chloride Test. The Colloidal Gold Test.— The gold 
chloride test has proven of great value in the recognition of early general 
paresis and of some value in differentiating the various types of meningitis. 
It is the most sensitive indicator we have of pathological changes in the 



2 Am. Jour. Insan., July, 1907, vol. lxiv. 



686 CLINICAL DIAGNOSIS 

spinal fluid. The reagents required to make up the colloidal solution of 
gold are a i% solution of gold chloride, a 2% solution of potassium car- 
bonate, a 1% solution of oxalic acid and a 2.5% solution of formaldehyde. 
All water used to make up these solutions must be triply distilled. To make 
a liter of the solution, 10 c.c. of the 1% gold chloride solution, 7 c.c. of the 
2% potassium carbonate solution, 1.75 c.c. of the 1% oxalic acid solution 
and 0.83 c.c. of the 2.5% formaldehyde are added to a liter of triply distilled 
water in a chemically clean flask. After thorough mixing the_fluid is heated 
to from 8o° to 85 C. and kept at that temperature until a series of color 
changes has taken place, which runs through gradations from faint blue-green 
to a deep ruby-red. When the solution reaches its maximum depth of 
color a remarkable lightening in hue occurs within the space of a few seconds, 
the dark ruby-red becoming converted to a lighter shade, and when this 
stage is reached the reaction is finished. The solution if well made has a 
ruby-red color, is transparent, neutral in reaction on the day on which it is 
used, and 5 c.c. of it will be completely precipitated in 1 hour by 1.7 c.c. 
of a 1% sodium chloride solution, which shows that it is not ' 'protected" 
from precipitation by electrolytes or by impurities. In addition, it must give 
characteristic results with known normal, paretic and luetic spinal fluids. 

It will usually keep indefinitely without spoiling, especially if protected 
from the sunlight. All glassware used in making and keeping this solution 
and in making the test must be perfectly clean, the glass itself neutral in 
reaction and the pipettes accurately graduated. The vessels of glassware 
maybe effectively cleaned by scrubbing them thoroughly with soap and hot 
water, rinsing for several minutes in running tap water, then immersing 
them in, or filling them with, the following bichromate-sulphuric acid mix- 
ture for at least one-half hour. 

Potassium bichromate, powdered 200.0 

Water, distilled, up to 1500.0 

Sulphuric acii, concentrated 500.0 

When ready to use the cleaning fluid is emptied out, the utensils are rinsed 
well in running water, then with distilled water, and finally they are flushed 
with triply-distilled water. 

It is also necessary to use pure water, which can be obtained by distilling 
it three times. This should be used as soon as possible after the third 
distillation because upon long standing it becomes unfit for use in making 
the gold solution. There should be no rubber connections on the still. 

In using the pipettes one should not blow the fluid out since even the 
carbon dioxide of the breath will disturb the results. This test is a practical 
application of the observation by Zsigmondy that proteins in general, each 
to a specific quantitative degree, protect the precipitation by a sodium 
chloride solution of a colloidal gold suspension. In the case of pathological 
spinal fluids certain proteins (albumins ?) may be present which precipitate 






CEREBROSPINAL FLUID 687 

the suspension while others (globulins?) protect the suspension against 
sodium chloride and against these other proteins. 

The effect of pathological spinal fluids on the suspension will depend on 
the character and relative proportion of the proteids and of other substances 
as well. What these proteins and "other bodies" are we do not know. The 
results are empirical and yet very valuable. In general paresis the sus- 
pension will be completely decolorized by a spinal fluid which gives none of 
the ordinary chemical tests for protein. 

Normal cerebrospinal fluid diluted even 1:5120 will prevent a 0.4% 
sodium chloride solution from affecting a colloidal gold suspension. The 
degree of change which the sodium chloride solution produces in the collodial 
gold suspension is indicated by the degree of discolorization of the suspension. 

The test is made as follows : A series of 1 1 large test tubes are arranged 
in a rack and numbered. Into the first tube are measured 0.2 c.c. of cere- 
brospinal fluid and 1.8 c.c. of a 0.4% solution of sodium chloride. Into 
each of the other tubes is measured 1 c.c. of the sodium chloride solu- 
tion. The contents of the first tube are now thoroughly mixed and 1 c.c. 
measured into tube 2. The contents of tube 2 are now thoroughly mixed 
and 1 c.c. measured into tube 3, and so on throughout the series until we 
come to tube 10 and 1 c.c. of the contents of this is discarded. Each of the 
10 tubes will now contain exactly 1 c.c. of a mixture in which the spinal 
fluid in tube 1 is diluted 1:10, tube 2 1:20, tube 3 1:40, etc., and tube 10 
1 :512c The eleventh tube will contain no spinal fluid and serves as a salt- 
solution color-control. One now adds to the contents of each tube 5 c.c. 
of the colloidal gold solution. The rack of tubes is set aside and observed 
from time to time. The reaction may appear in an hour but the final 
reading should not be made until the end of 24 hours. The tubes are to 
be examined by direct daylight, not by artificial light. 

The change in color from the original ruby-red (o) due to different de- 
grees of change in the gold suspension are red blue (1), violet (2), blue (3), 
gray (4) and colorless (5). 

The typical reactions are: 

Normal = 00000, 00000 

Luetic = 01233, 31000 

Non~Luetic = 00012, 34210 

Paretic =55555, 42100 

These reactions may be expressed graphically as curves as in Fig. 144, 
in which A is the curve of normal fluid, B the non-luetic curve, C the luetic 
and D the paretic curve. 

That part of the chart represented by the dilutions 1:10 to 1:160 has 
been called the luetic zone, and that from 1 :i6o to 1 :i28o the non-luetic or 
meningeal zone. The paretic curve is the most constant of all. This in 
incipient paresis may be positive when the spinal fluid and blood Wasser- 
manns are negative, the cell count within normal limits and the test for 



688 CLINICAL DIAGNOSIS 

globulin negative. In the diagnosis of this disease the test exhibits its 
greatest diagnostic value. The luetic curve is sometimes positive in cases of 
tertiary lues without any symptoms or other evidence of luetic involvement 
of the central nervous system. The maximal color change in cases of tuber- 
culous and epidemic cerebrospinal meningitis always occurs in the higher 
dilutions to the right of the midline. This test is of no aid in a differential 
diagnosis between the various forms of non-luetic meningitis, the tuber- 
culous and suppurative forms. 

It is of great advantage that spinal fluids to be examined by this method 
need not be particularly fresh. 

CEREBROSPINAL FLUID IN DISEASES 

Uremia. — In cases of nephritis showing the syndrome called uremia, 
and especially those with convulsions, the cerebrospinal fluid is greatly 
increased in amount and therefore on puncture will spurt from the needle in 
a steady stream under considerable pressure. Even 40 c.c. or more may 
escape before the pressure reaches normal. 

The cell count may or may not be increased. The chlorides are increased 
even to 0.85 gms. per 100 c.c. The urea is greatly increased in amount. 
Lactic acid has been demonstrated. 

Diabetes Mellitus. — In diabetes mellitus the cerebrospinal fluid has 
been found normal in appearance, amount, pressure, cell count and in every 
particular chemically except that glucose is increased. Foster found this 
even 3%. Levinson's highest figure was 0.38%. In one case this latter 
author found it higher in this fluid than in the blood. Acetone and diacetic 
acid may be present. 

Chorea. — In Sydenham's chorea there is sometimes hypertension and 
a distinct lymphocytosis of the spinal fluid, but most observers find this 
fluid normal. 

Epilepsy. — During a convulsion of true idiopathic epilepsy the pressure 
of the cerebrospinal fluid is increased. Between attacks it is normal chem- 
ically and in pressure. 

Mental Diseases. — In the psychoses not dependent on some demon- 
strable disease, as lues, the cerebrospinal fluid is normal in every particular. 
In alcoholic psychoses the fluid is increased in amount and pressure while 
the cell count is as a rule normal. 

In hydrocephalus the fluid, while increased in amount, is practically 
normal in every other way. 

In cases with recent cerebral hemorrhage the cerebrospinal fluid is red 
. from the presence of blood and later yellow from the presence of modified 
hemoglobin. The cell count and proteid content will depend on the amount 
of blood present. 

In tumors of the brain the fluid may spurt through the needle under 



CEREBROSPINAL FLUID 689 

high pressure. In such a case the flow should be checked at once and not 
over 5 c.c. in all removed, for the higher pressure in the cranial cavity when 
that in the spinal canal becomes lower may crowd the medulla into the 
foramen magnum and cause death. When the tumor is so located that it 
causes stasis of the flow of this fluid its protein content and cell count may 
be much increased. 

Compression of the Cord. — In tumors of the cauda equina and 
conus medullaris the spinal fluid often shows a yellowish color (xantho- 
chromia), its proteid is greatly increased and a lymphocytosis is present 
(Froin's syndrome). 

If the tumor is at a higher level there may be no lymphocytosis but an 
increased amount of globulin and usually a xanthochromia. 

Acute Encephalitis. — Excluding that which is a part of meningitis or 
poliomyelitis and that due to the acute infectious diseases, encephalitis is a 
condition easy to assume but difficult to define. In those cases with clinical 
symptoms suggesting such a condition the spinal fluid usually is normal or 
slightly increased in amount and the cell count perhaps a little increased, 
but otherwise it is normal. In some cases of epidemic encephalitis a slight 
increase in the cell count (from 10 to ioo) and a slight increase in globulin 
was reported but in other cases this fluid was reported normal. 

Meningism. — The term meningism is applied to that large group of 
cases, most of them of the acute infectious fevers in children, with many 
clinical symptoms of meningitis but with a spinal fluid sterile and clear, 
usually with normal cell count and normal globulin, but sometimes under 
increased pressure. 

Epidemic Cerebrospinal Meningitis. — In epidemic cerebrospinal men- 
ingitis, the spotted fever and epidemic cephalalgia of older writers, the 
spinal fluid is increased in amount and pressure (unless the exudate has be- 
come so thick, literally buttering the cord, that none could possibly flow 
through a needle. It is of interest that even such an exudate, demonstrated 
by laminectomy, may disappear entirely, leaving the meninges quite free 
from any evidence of previous inflammation.) The pressure of the fluid 
is as a rule increased from 300 to even 800 mm. of water. Its color may vary 
from a slight greenish turbidity to that of cream. On standing the pus by 
settling forms a sediment of varying amount. 

The protein content is greatly increased, even to 0.7%, and the sugar is 
greatly diminished. The cell count may reach several thousand per c.mm. 
the great majority of which (90 to 98%) are polymorphonuclear finely 
granular cells. In the smears may usually be found a few, minute, Gram- 
negative, biscuit-shaped diplococci, the most of them intracellular. This 
memingococcus grows on most culture media and in that way differs from 
other diplococci of similar morphology which also are Gram-negative. 

H 



690 CLINICAL DIAGNOSIS 

From some cases both the meningococcus and pneumococcus may 
be isolated. 

Micrococcus Intracellulars Meningitidis. — This organism (Fig 
145) closely resembles the gonococcus in morphology. While some are 
extracellular, its most typical appearance is as intracellular biscuit-shaped 
diplococcus which varies noticeably in size. The meningococcus is Gram- 
negative, is stained easily with the common bacterial stains and can be 
grown on many of the common culture media. 

Pneumococcus Meningitis. — In pneumococcus meningitis the fluid 
escapes under high pressure, is milky- white as a rule and has not the greenish 
color almost constant in cases of the epidemic form, than which also it is 
more purulent, is richer in fibrin and deposits a more abundant sediment. 
The cell count is high, the leucocytes chiefly polymorphonuclear finely 
granular cells and the organism (Micrococcus pneumoniae) more numerous 
in the smears and easily grown in cultures (Fig. 146). 

Streptococcus Meningitis. — The fluid in streptococcus meningitis 
resembles that due to Micrococcus pneumoniae except that streptococci 
can be demonstrated in smears and in cultures. Both the hemolytic and 
non-hemolytic forms may be found. 

Influenza Meningitis. — The fluid is a very thin pus containing 
Bacillus influenzae in great numbers. (Fig. 147). 

Typhoid Meningitis. — F. M. male, aged 20, No. 5 146, was admitted August 2, 191 7 
and died August 7, 19 17. On admission he was clearly typhoid in condition, evidently 
in the second week of this disease, with apparently a meningeal involvement. August 
3rd the spinal fluid was negative in appearance and on examination except that it gave 
a one plus globulin test and had a cell count of 6 per c.mm. On August 4th the cell 
count was 15 and the culture of the fluid was positive for Bacillus typhosus. On this 
day also the blood Widal was positive and the leucocyte blood count 7600. August 5th 
Bacillus typhosus was found in both spinal fluid and blood cultures. August 7th . the 
blood leucocyte count was 36,600 and the spinal fluid purulent. Stained smears of the 
spinal fluid showed it to contain many bacilli. 

Tuberculous Meningitis. — In tuberculous meningitis, the acute hy- 
drocephalus of former writers, the fluid early is under greatly increased 
pressure, from 300 to 700 mm. of water (from 240 to 550 mm.; average 
365 mm.) 3 but this slowly decreases as coma develops. It usually is quite 
normal in appearance when drawn, is crystal clear even though it may con- 
tain even 500 cells per c.mm., but it develops a foam when shaken and 
on standing a fine web-like clot of fibrin appears which alone is almost 
sufficient for diagnosis. The specific gravity is as a rule about 1 .006. There 
is a definite lymphocytosis of from 80 to 952 cells per c.mm. although early 
many of the cells may be polymorphonuclear finely granular leucocytes. 

3 Rous, Am. J. Med. Sc. 1907. 



CEREBROSPINAL FLUID 691 

It is of importance in diagnosis that in this disease the count usually 
is above ioo while in lues of the central nervous system it is usually below 
ioo. The protein is greatly increased in amount, ranging between o.i 
and 0.2 %; the permanganate index is above 2; the chlorides are re- 
duced, are usually below 0.6%; the glucose as a rule varies between 0.5 
and 0.6%; and the H-ion concentration of the fresh fluid averages from 
7.4 to 7.6. 

The colloidal gold test shows a discoloration of tubes 5-6-7-8 and gives 
a curve of which 00012332 10 is typical. 

For a positive diagnosis one must demonstrate Bacillus tuberculosis. 
The fresh fluid, diluted with an equal volume of alcohol to lower its specific 
gravity, may be centrifugalized vigorously and smears made from this sedi- 
ment. If a reticulum has formed this pellicle will contain the most of these 
organisms and smears made of this will often demonstrate the organism. 
Or, the- fresh fluid (or the fluid after any clot present has been digested) 
may be injected into a guinea pig. 

Syphilis. — In every case of syphilis, no matter how early, the spinal 
fluid should receive careful attention. There is indeed evidence that the 
so-called neurasthenia present in the secondary stage may indicate an early 
involvement of the nervous system. Certainly no case should be dis- 
charged from treatment as cured unless the Wassermann and cell count of 
the spinal fluid are negative as well as the Wassermann of the blood. Many 
relapses in cases clinically and seriologically cured are due to reinfection 
of the vascular system from the cerebrospinal system. 

Cerebrospinal lues, tertiary syphilis of the central nervous system, is 
accompanied by an increase in the amount of spinal fluid, an increase in 
the protein, in the cell count, by a positive Wassermann reaction and a 
fairly typical colloidal gold curve. 

In general paresis the examination of this fluid is most important. This 
will usually demonstrate an increase in amount, a lymphocytosis of moderate 
grade, an increase in the globulin, a typical colloidal gold curve and, in 90% 
of the cases, a positive Wassermann reaction. 

Acute Syphilitic Meningitis. — In acute syphilitic meningitis the 
fluid is slightly increased in amount and so is under slightly increased pres- 
sure, is clear or slightly opalescent, the globulin tests are positive, there is a 
definite pleocytosis, (the counts varying from 100 to 600 per c. mm., 60 to 
80% of which are lymphocytes), the Wassermann test is positive and the 
colloidal gold test gives a curve which is luetic in character. 

The fluid will be sterile on culture for bacteria but Spirochete pallida 
has been demonstrated in the sediment. 

This condition should be excluded in all cases with clinical symptoms 
suggesting menginitis and with fluid sterile on culture. 



692 



CLINICAL DIAGNOSIS 



Acute Anterior Poliomyelitis. — In poliomyelitis the spinal fluid is 
increased in pressure (from 300 to 700 mm. of water) and in amount, since 
from 20 to 50 c.c. are easily obtained on puncture. Usually it is colorless, 
but sometimes slightly opalescent, and foams considerably on shaking. The 
globulin is slightly increased as a rule but not always. 

The cell count is increased, the pleonucleosis manifesting itself in the 
pre-paralytic stage and lasting from 14 to 16 days after the appearance of 
the paralysis. The count is highest during the first week of the paralysis. 
Very early the polynuclear cells may predominate, but later the percentage 
of small mononuclears will vary from 60 to 90%. The spinal fluid is sterile 
by ordinary cultural methods, while some authors have found a coccus which 
grows best under aerobic conditions on a 1% glucose broth medium. (Nu- 
zum, Rosenow) . Flexner and Lewis found a filterable virus. 



CHAPTER VII 

EXAMINATION OF VARIOUS FLUIDS 

Among the various fluids which may deserve the attention of the 
diagnostician are the plasma, serum, lymph, the various transudates and 
exudates, the cystic fluids, the synovial and the amniotic fluids. 

Specific Gravity. — The specific gravity of a fluid may be determined 
with an accurate aerometer (see page 429) if there is sufficient quantity at 
hand but more accurately gravimetrically. The figures given in the fol- 
lowing pages were determined by this latter method. 

Dried Constituents. — The dried constituents of a fluid are determined 
by measuring or weighing into a weighed glass dish with a ground glass 
stopper from 10 to 30 c.c. of the fluid in question. This is evaporated over 
a water-bath and then in vacuo over sulphuric acid. It is then dried to con- 
stant weight at temperature not over no° since urea or other fragile bodies 
usually are present and would be broken down by the heat. 

Proteids. — Among the proteids which may be met with in body fluids 
are serumalbumin, serumglobulin and, in some, fibrinogen. The albumoses 
are rare. True peptone is said never to occur, while the glyco-proteids and 
the phospho-proteids may be met with, e. g., in the cystic fluids. To isolate 
the proteids it is first necessary to remove any organized structures. This 
may be done by sedimentation, centrifugalization or by filtration through 
paper or Kieselguhr. Fibrin will be evident to the naked eye, will disappear 
rapidly in artificial gastric juice, and undergoes a glassy swelling on the 
addition of 0.1% HCL 

Albumin, Globulin, Fibrinogen. — From 20 to 50 c.c. of the fluid 
are mixed with an equal amount of saturated (NH 4 ) 2 S0 4 and allowed to 
stand 1 hour. The amount chosen should not contain more than 0.2 to 0.3 
gm. of proteid for each precipitate. It is then filtered through a weighed 
filter and the precipitate washed with half -saturated (NH 4 ) 2 S0 4 until the 
filtrate gives no cloud with acetic acid and K 4 FeCN 6 . 

(a) The filtrate is boiled, acetic acid added until it is faintly acid, then 
boiled again and filtered through a weighed ashless filter. The precipitate is 
then washed with hot water, then with alcohol, then with ether and 
brought to a constant weight at 120 C. It is then ashed and the weight 
of this subtracted. 

(b) The precipitate on the filter paper is heated to 1 io°, washed with hot 

water, then with alcohol and ether, dried to constant weight and its ash 

subtracted. This will equal the weight of the serum globulin and fibrinogen. 

For the determination of both together see page 698. 

693 



694 CLINICAL DIAGNOSIS 

Glyco-Proteids and Phosphorus-Containing Proteids. A. Mucin. — In 

general, to isolate mucin ioo c.c. of fluid are diluted if necessary with water, 
precipitated with acetic acid, filtered and the precipitate washed with water 
acidulated with acetic acid. The precipitate is then dissolved in weak al- 
kaline water and reprecipitated with acetic acid. 

A part of this precipitate is boiled on the water -bath with dilute mineral 
acid (HC1), filtered and the filtrate tested for sugar to demonstrate the 
reduction of copper. Considerable boiling may be necessary and the re- 
duced copper may be seen only after the fluid is cold and the precipitate 
has settled. 

Mucin is a glyco-proteid of a stringy consistency and insoluble in 
acetic acid even in excess. Mucoid is similar in nature, but differs in some 
physical characteristics. A sharp line cannot be drawn (see page 219). 

B. Phosphorus-containing Proteid. — A part of the precipitate is 
examined for organic phosphorus. It is ashed, the ash dissolved in dilute 
HNO3, heated to boiling, concentrated somewhat and then ammonium 
molybdate added in excess. A yellow color and then a yellow precipitate 
which forms most readily at 40 C. is evidence of phosphoric acid. Or, 
the precipitate may be dissolved in HC1, made strongly alkaline with 
ammonia and then magnesium mixture added. The white precipitate of 
NH 4 MgPO 4 indicates the presence of phosphoric acid. 

If the reaction for phosphorus is faint the test has no meaning since all 
of the phosphate and lecithin cannot be washed from the mucin precipitate. 

If organic phosphorus has been found, a part of the original precipitate 
is dissolved in NaOH, then HC1 added, boiled to clear solution, super- 
saturated with ammonia and then precipitated with AgN0 3 . A flocculent 
cloud of the silver salt of the nuclein base indicates a nucleo-proteid. If 
none forms, the phosphorus body is a paranucleo-proteid. 

Fat, Lecithin and Cholesterin. — To from 20 to 50 c.c, weighed or 
measured, of the fluid to be examined for fat, lecithin and cholesterin are 
added from 3 to 4 volumes of absolute alcohol. The specimen is then 
allowed to stand until the next day during which time it is repeatedly 
stirred. It is then filtered and the precipitate washed with absolute al- 
cohol and placed in the cylinder of a Soxhlet ether-extraction apparatus. 
The alcohol filtrate is now neutralized and evaporated at 6o° C. The residue 
is taken up with alcohol and ether and re-evaporated. The residue is then 
taken up with ether and poured into the flask of this same Soxhlet appa- 
ratus. The precipitate is then extracted for hours. The ether extract is 
evaporated, the residue taken up in water-free ether, the filtered solution 
evaporated in a weighed beaker "and dried to constant weight in vacuo over 
sulphuric acid. This residue is made up of fat, lecithin and cholesterin. 

The residue is now dissolved in alcohol, alcoholic KOH added, warmed 
on the water-bath for one hour and then evaporated to dryness. The fat 



EXAMINATION OF VARIOUS FLUIDS 695 

is now in the form of soap and glycerin. To it is now added water (not 
too little) and it is shaken out several times with equal volumes of ether. 

This ethereal extract is distilled to small volume, then evaporated in a 
weighed beaker to dryness. The residue contains soap and cholesterin. 
The soap may be washed out with small portions of cold alcohol slightly 
acidulated with HC1. The cholesterin left is dried at 8o° C. and weighed. 

The alcohol washings with the soap are added to the water extract of the 
previous separation which now contains all the lecithin-phosphorus. This 
fluid is evaporated, the residue ashed and its phosphorus determined. 

(Distearyllecithin contains 3.84% of phosphorus and dipalmityl- 
lecithin, 4.12%.) 

Leucin and Tyrosin. — A fluid to be examined for leucin and tyrosin is 
worked with as fresh as possible. All albumin should be removed by heat 
and acetic acid, or by precipitation with from 3 to 4 volumes of alcohol, 
then heated on the water-bath, cooled and filtered. The alcohol is then 
removed by evaporation. The nitrate is precipitated with neutral, then 
with basic, lead acetate, avoiding carefully any excess, and filtered. The 
lead is removed from the filtrate with H 2 S, the filtrate then evaporated 
and examined for crystals (see page 254). 

Succinic Acid, CH 2 COOH.CH 2 COOH .— Traces of this acid are met 
with in many animal fluids; sometimes in the fluid of hydrocephalus and 
hydrocele, in larger amounts in echinococcus cysts and in wool-fat. It is 
formed by the bacterial decomposition of proteids and sugar. It frequently 
occurs in acid milk in the intestine, in putrid pus and whenever alcoholic 
fermentation of sugar is in progress. 

Any fluid to be examined for succinic acid is freed of albumin by heat 
and acetic acid. (If it be urine which is tested the albumin is first removed, 
the urine perfectly precipitated with baryta water and the excess of this 
removed by H 2 S0 4 .) The filtrate is next evaporated to a residue, acidified 
with HC1 and extracted repeatedly with ether. The ether is evaporated off, 
the residue taken up with a small amount of water and allowed to stand 
until crystallization. Or, the watery filtrate may be heated to boiling, 
nitric acid added drop by drop until it takes a slight yellow color and then 
evaporated . If no crystals form a portion of the residue is fused in a test- 
tube with ammonia and zinc dust. If a match-stick wet with strong sul- 
phuric acid is held at the mouth of the tube the red color of pyrrol would 
indicate succinic acid. If this test is used, however, hemin and the indol- 
derivatives must be excluded. These latter will give the reaction on heating 
alone and hemin on heating with zinc dust alone. 

Lactic Acid, C 3 H 6 3 . — Of the 3 modifications of lactic acid the inactive, 
or the lactic acid of fermentation, occurs oftenest in the stomach and in- 
testine of man. The dextro-rotatory form, or sarcolactic acid, may be 
demonstrated in the muscles, blood, pericardial fluid, aqueous humor and 
intestinal contents. It occurs also in the urine in acute yellow atrophy and 



696 



CLINICAL DIAGNOSIS 



phosphorus poisoning, liver cirrhosis, after respiratory distress, severe 
exercise and before death. It occurs in pathological transudates often in 
abundance, in the bones in osteomalacia and in the sweat in puerperal fever. 
The levo-rotatory form has never been found in the body. 

The fluid to be examined for lactic acid is, if necessary, made faintly 
acid with dilute H 2 SO 4 and is boiled and filtered to remove any albumin. 
Baryta water is added to the filtrate as long as a precipitate forms and the 
excess of the barium removed with C0 2 . The nitrate is now evaporated to 
a thin syrup without heating it above 70 , in order to avoid the develop- 
ment of a brown color. About ten volumes or more of absolute alcohol 
are then added slowly to the syrup, this well stirred, allowed to stand for 
some time and the alcohol then poured off. The residue is dissolved in a 
little water and this procedure is repeated once more with alcohol. 

The combined alcoholic solution is poured off, filtered, the alcohol 
distilled off, leaving a thin syrup which is digested on the water-bath at a 
moderate temperature to drive off the alcohol and then cooled. To the 
resulting thin syrup is added an equal amount of dilute phosphoric acid. 
It is then brought into a large flask and shaken out with a large amount of 
ether which will gradually take up the lactic acid. The ether must be fre- 
quently renewed. The united ether extracts are then filtered clear, the 
ether distilled off, the residue dissolved in water, boiled for some time with 
an excess of ZnC0 3 , filtered, washed with hot water, evaporated to a small 
volume on the water-bath and allowed to stand until the zinc salt of lactic 
acid crystallizes out. Alcohol is then added to the mother liquid which is 
then allowed to stand longer and another mass of crystals is obtained. The 
zinc salt is dissolved in hot water and the zinc precipitated by H 2 S. The 
filtrate is then evaporated to a syrup containing the lactic acid. 

It is quite impossible to recognize lactic acid from its crystalline form 
or by Uffelmann's test alone. 

Inosite C 6 H 6 (OH) 6 . — Inosite is found in traces in the urine of perhaps 
every normal person. It certainly is present in cases with polyuria and 
therefore in the urine of some patients with diabetes and nephritis. It is 
found also in the contents of echinococcus cysts. 

To demonstrate inosite the fluid is freed from albumin by heat and acetic 
acid, the phosphates are precipitated by baryta water, the filtrate is evapo- 
rated and the creatinin allowed to crystallize out by boiling with from one 
to four volumes of alcohol. If a heavy precipitate results which sticks to the 
glass the fluid is simply decanted, but if flocculent, it is filtered through a 
heated filter and then allowed to cool. The fluid then stands for 24 hours. 
If inosite is present crystals will form which may be filtered out and washed 
with cold alcohol. The alcohol precipitate mentioned above may be dis- 
solved in boiling water, from 3 to 4 volumes of hot alcohol added and the 
above procedure repeated to recover the inosite therein contained. If no 
crystals form one adds ether, little by little, to the clear alcoholic filtrate 



EXAMINATION OF VARIOUS FLUIDS 697 

until a slight milky cloudiness results which does not disappear. The speci- 
men is then allowed to stand for 24 hours. All the inosite will be precipitated 
as mother-of-pearl plates. In case urine is the fluid examined, one first 
precipitates with baryta water and precipitates the filtrate, after heating, 
with PbAc, avoiding an excess. The specimen is then allowed to stand, 
is filtered, the precipitate washed, suspended in water, decomposed by 
H 2 S and the filtrate evaporated. One then proceeds as above. 

Inosite crystallizes in rhombohedral crystals which melt at 225 C, 
are soluble in 1 .75 of water and soluble in alcohol or ether. It does not 
ferment, nor does it rotate the plane of polarization; it dissolves Cu(OH) 2 
without reduction, is precipitated by PbAc and does not give crystals with 
phenylhydrazine . 

Scherer's Test. — A small amount of the precipitate is evaporated almost 
to dryness with nitric acid on a platinum-foil. To the residue are added am- 
monia and 1 drop of CaCl 2 and the evaporation then continued to dryness. 
A beautiful rose color is the result. The crystals must be quite pure to 
give a positive test. 

SeideVs Test. — This test is similar to the above with the exception that 
strontium acetate is used instead of calcium chloride and the positive re- 
sult is a green color with a violet precipitate. This test is positive if o. 3 mgm. 
of inosite is present. 

Allantoin, C4H6N4O3. — This body is found in the urine of the new-born 
child. A slight trace is said to occur in all normal urines and especially 
that of pregnant women. It occurs in some ascitic fluids, as in liver 
cirrhosis and in certain ovarian cysts. 

To demonstrate allantoin the albumin of the fluid to be examined is 
removed by heat and acetic acid. The fluid is then precipitated with 
Hg(N0 3 ) 2 , the precipitate washed, suspended in water, decomposed with 
H 2 S and filtered. A little ammonia is added to the filtrate and the whole 
evaporated on the water-bath to a small volume. The clear fluid is then 
precipitated with ammoniacal AgN0 3 . (Since the precipitate is soluble 
in an excess of ammonia this must be avoided.) After the specimen has 
stood for some time the silver salt of allantoin is collected on the filter, is 
washed, suspended in water, decomposed with H 2 S, the filtrate evaporated 
and allowed to crystallize. 

The Loewy method is recommended for the demonstration of allantoin 
in the urine. The faintly acid urine is precipitated with mercurous nitrate 
(which is dissolved in as little acid as possible plus some metallic mercury) , 
is filtered and the precipitate well washed. The filtrate is then precipitated 
with H 2 S and filtered, the filtrate warmed to drive off this gas, MgO then 
added and the whole precipitated with AgN0 3 . This precipitate is filtered, 
washed, suspended in water and decomposed with H 2 S while warm. This 
filtrate is evaporated to dryness, the residue extracted with hot water and 
when cold precipitated with Hg(N0 3 ) 2 . This precipitate is well washed, 



698 CLINICAL DIAGNOSIS 

decomposed with H 2 S, the filtrate evaporated to a concentrated solution 
whereupon the allantoin will crystallize out in glistening prisms, which are 
odorl'ess, tasteless, soluble in 160 parts of cold water and more in warm, and 
insoluble in absolute alcohol or ether. For its identification the silver salts 
are studied, which are obtained by precipitating a concentrated solution by 
ammoniacal AgN0 3 . This precipitate is soluble in excess of ammonia. 
The white flocculent precipitate on standing becomes granular. If dried at 
ioo° it gives an easy reduction of silver. The silver salt of the allantoin 
dried in vacuo gives on fusion 40.71% of silver. 

Quantitative Analysis of Serous Fluids. — The method given by . Thierfelder 1 
is the one we have used. It is fairly satisfactory as a routine preliminary method 
for getting an idea of the general composition of a fluid, but this determined it is 
better to determine each substance or group in a special portion. We give the outline 
of the analysis. 

To from 20 to 50 c.c. of the fluid, measured or weighed and freed from the formed 
elements by filtration, are added from 3 to 4 volumes of absolute alcohol. This is allowed 
to stand a few hours, filtered through a weighed filter and washed a little with alcohol. 
This will give Filtrate I. The precipitate on the filter paper is then washed with boiling 
alcohol, again and into the same flask with ether and then again with alcohol. This 
gives Filtrate II. The precipitate is now washed with boiling water thoroughly. This 
gives Filtrate III. The precipitate remaining on the paper will contain the proteid, a 
few salts, hemoglobin if any be present, but not much of the other pigments. It is now 
washed once more with alcohol and after the precipitate next to be mentioned has been 
added to it dried at 120 C. to constant weight. It is then ashed and the weight of the 
ash subtracted from that of the precipitate; this gives the weight of the proteids. 

Filtrate I is evaporated at a temperature not above 6o° C. To this residue is then 
added Filtrate II. The residue and fluid are well mixed and the fluid decanted through 
a dried and weighed paper. It is then washed repeatedly with absolute alcohol, then 
with ether and decanted each time, but none of the precipitate is allowed to get onto 
the paper (B 3). Onto the residue is now poured Filtrate III, the residue is well mixed 
and then all filtered through the same filter-paper, but into another flask from that which 
has received the above mentioned decanted fluids.- The precipitate is now brought on 
the paper and washed with water. This precipitate is dried and added to the above 
mentioned proteid precipitate, since it contains that amount of proteid which was lost 
in the first precipitation. The watery extract of the above is evaporated in a small 
weighed porcelain dish in a water-bath, dried at no° to 115 C. to constant weight 
and washed. 

B 2 is burned at a moderate heat, ashed, and the weight of the ash determined. 

B 3. This alcohol-ether extract is evaporated on the water-bath at a temperature 
not above 6o° C. and dried in vacuo over H^SO-i- To the residue is added ether and it 
is filtered into a flask through a small paper, washing repeatedly with ether. This will 
contain the urea, sugar, soaps, sodium chloride, fat, lecithin, cholesterin and the choles- 
terin ester. 

B 3a is the residue of the above. This is washed from the beaker and paper into a 
small weighed porcelain dish, evaporated, dried at a temperature from no° to 115°, 
fused at a moderate heat and weighed. 

B 3&. This ethereal extract is treated as on page 557. It contains fat, cholesterol, 
lecithin, cholin, etc. 

x Hoppe Ssyler, Chemische Analyse, 1903. 



EXAMINATION OF VARIOUS FLUIDS 699 

The Fat Percentage of Milk may be determined by mixing well in a 
large Babcock centrifuge tube 

15.0 c.c. of well mixed milk 
1.5 c.c. of amyl alcohol and 
1.5 c.c. of cone. HCL 

and then added 10 c.c. of concentrated H 2 S0 4 . The specimen is again well 
mixed and the Babcock tube rilled to the upper mark with 50% H 2 S0 4 . 
It is centrifuged for 3 or 4 minutes and then the fat percentage read off. 

To determine the milk proteids one estimates the total nitrogen (see 
page 107) and multiplies by the factor 6.37. 

To count the bacteria in milk one dilutes the milk 1:10 and 1:100 and 
plates with agar-agar using from 0.1 to 1.0 c.c. of the diluted milk. The 
plates are kept for 1 night in the incubator at 3 7 C. and the colonies counted 
on the following day. 

TRANSUDATES AND EXUDATES 

Although the pathologists believe that no sharp distinction should be 
attempted between exudates and transudates so far as the mechanism of 
their origin is concerned, yet the clinical chemist is forced to differ entiate 
between them on the basis of their physical and chemical properties. In 
some measure at least a transudate is less the result of an inflammation than 
is an exudate. The transudates resemble lymph; they contain few formed 
elements and almost no fibrin. Their proteids are serum albumin, serum 
globulin and a little fibrinogen, yet not enough of the last to coagulate these 
fluids spontaneously, although they will coagulate if blood is added. The 
exudates are richer in formed elements, coagulate spontaneously, contain 
the so-called "nucleo -albumin" and a mucoid substance. Some claim that 
fluids whose specific gravity is under 1.018 are usually transudates; and 
those in which it is over 1.018, are exudates. The list of extractives 
which may be present in these fluids includes urea, glucose, creatinin, 
uric acid, lactic acid, inosite, succinic acid and allantoin. More rarely one 
finds leucin, tyrosin, bile acids and pigments, fat, lecithin and cholsterin. 

The presence of "nucleo-albumin" or, better "euglobulin," is valuable 
in distinguishing these two classes of fluids. If a few drops of acetic acid 
are added to a clear exudate a cloud of varying depth, but usually quite 
dense, will form. It is rather soluble in excess of the acid. The cloud in 
transudates is very much lighter. 

Peritoneal Fluid. — In cachexia and hydremia this fluid is slightly 
colored, of a milky opalescence, does not clot spontaneously, has a specific 
.gravity of from 1.005 to I - OI 5 an d contains almost no cells. 

In chronic passive congestion the specific gravity is usually lower 
than 1.020. Sometimes it contains 35 gms. per liter of proteid. Incases 
of cancer of the peritoneum the fluid is turbid with cells, has a dirty grayish 



700 CLINICAL DIAGNOSIS 

appearance, a high specific gravity and often clots spontaneously. The- 
serous fluid present in inflammations is of a straw or lemon-yellow color, 
is somewhat cloudy from the formed elements, coagulates spontaneously, 
contains 30 gms. or more per liter of proteid and has a specific gravity of 
1.030 or above. Mucoid substance is perhaps always present, which may- 
be demonstrated by removing the albumin by heat and then precipi- 
tating the nitrate by alcohol. A precipitate forms from which one can. 
split off a reducing body. 

In the ascitic fluid may also be determined urea, uric acid, allantoin,. 
xanthin, creatinin, cholesterin and sugar. 

The ascitic fluids we have examined (none were very acute cases) had a. 
specific gravity varying from 1.0055 to 1. 0198, "the solids from 1.3 to 4.5 gms. 
per liter and globulin 40 to 50% of the proteid. 

Pleural Fluid. — Physiologically there is not enough pleural fluid present 
for an analysis. The pathological fluid may be of any degree of serous, 
sero-purulent, purulent or hemorrhagic quality. In hydrothorax the specific: 
gravity is lower than 1.015 as a rule, the albumin from 10 to 30 gms. and. 
the fibrinogen hardly 0.1 gms. per liter. In pleurisy the exudate has a 
specific gravity above 1.020 as a rule, albumin 30 to 65 gms. and the^ 
fibrinogen 1 gm.'per liter. 

In 9 of our cases the specific gravity of the fluids varied from 1. 01 22 to 1.0252 and 
the solids from 3.12 to 7.926%. The more acute the case the higher the figures. The- 
amount of total proteid varied from 2.837 to 6.529%, of which from 39 to 64% was 
globulin. The amount of globulin depended on the acuteness of the case. The acetic- 
acid precipitate was markedly more in the acute inflammatory cases than in the transu- 
dates. It is interesting what little difference the clotting makes. In a case of acute- 
tuberculous pleurisy before the fluid clotted its specific gravity was 1.02 21 and the 
globulin 2.875%; after clotting (densely) these figures were 1.02 17 and 2.376% respec- 
tively. To clear these fluids with the centrifuge is better than with Kieselguhr. 

In typical cases of pleurisy with effusion, and these are usually tuber- 
culous in character, the fluid is yellow or faintly greenish-yellow in color, 
with a specific gravity of at least 1.018 and an albumin content of 4% or 
more. On the addition of acetic acid an abundant precipitate falls. The 
fluid contains much fibrinogen and therefore coagulates rapidly, forming- 
some times small clots, or the entire mass of fluid may become a gelatinous 
clot (rarely must a little blood be added to aid clotting) . Microscopically 
it contains leucocytes and a few red blood-cells. In the majority of cases the 
most of the leucocytes are small mononuclears. 

Cytodiagnosis is the diagnosis of the etiology of an effusion based on 
the formula of its cell count. The fluid to be examined should be received 
into a sterile flask about % full of 0.85% sodium chloride solution and con- 
taining also 1% sodium citrate. This fluid is centrifugalized and the smears, 
made from the sediment are stained in the same manner as are blood smears. 
One can recognize in these smears the following leucocytes (Fig. 148) : small. 



EXAMINATION OF VARIOUS FLUIDS 701 

mononuclears, polymorphonuclear finely granular cells, polymorphonuclear 
coarsely granular cells, basophiles and endothelial cells. The latter de- 
mand special attention. Some may be the large mononuclear leucocytes 
of the blood (the so-called endothelial leucocytes) while others occur in 
sheets as though stripped directly from the pleural membrane. Some are 
phagocytic. All of these leucocytes are apt to have pycnotic nuclei and a 
vacuolated, degenerated protoplasm. Widal claims that if the majority 
of the cells in an exudate are finely granular the case is one of pyogenic 
infection; if the small mononuclears, it is one of tuberculous pleurisy, while 
in pleurisies of mechanical origin the endothelial leucocytes will predomi- 
nate. But the formula varies not only with the etiology but also with the 
intensity of the infection and with the duration of the effusion. We may 
have a transitory leucocytosis in cases of early tuberculous pleurisy ; we may 
also in such a case have a secondary infection of the pleura with pyogenic 
organisms. Many effusions which are clearly non-tuberculous in character 
may later have a preponderance of small round cells in the fluid. In fluids 
due to mechanical causes the relative excess of endothelial cells is early 
and transitory. Later, the small mononuclears may predominate. The 
polymorphonuclear finely granular cells seem least subject to variation. 
Some cases with pleural effusion in which the small mononuclear leucocytes 
are clearly in predominance are at autopsy definitely non-tuberculous. 
In cases of malignant disease involving the pleura the fluid may at first 
resemble that due to mechanical causes, but in other cases the mulberry- 
like masses of proliferated endothelial cells (proliferative pleurisy) and the 
cells showing mitotic figures may aid in the diagnosis, while in some cases 
of sarcoma involving the pleura the numbers of spindle-shaped cells may 
suggest the cause. Lord, in 28 cases of pleurisy with effusion, found the 
small mononuclears from 90% to 100% in 24; 88% in 1; and from 70% to 
75% in 3 cases. In the last three the epithelial leucocytes and the coarsely 
granular cells were prominent. If the fluid is complicated by pulmonary 
tuberculosis we may have a mixed infection and therefore a larger number 
of polymorphonuclear cells. In chronic passive congestion also there is a 
lymphocytosis; therefore a predominance of these cells does not always 
indicate tuberculosis, although it usually does. In cases of non -tuberculous 
pleurisy with clear serous effusion the polymorphonuclear neutrophiles may 
be in excess; in some, the small mononuclears predominate. The small 
mononuclears may predominate also in a mild pyogenic infection which is 
subsiding. In transudates the epithelial cells predominate, but if the fluid 
is of long standing, the small mononuclears. Secondary infection may con- 
fuse the picture. The fluid may also contain red blood-cells. Cholesterin, 
uric acid and sometimes sugar also are found. .In its chemistry the fluid 
resembles blood plasma. The proteids are serum-albumin, serum-globulin, 
some nucleo-albumin and fibrinogen. The albumin will be over 4% in 
amount. The fluids may coagulate spontaneously. 



702 CLINICAL DIAGNOSIS 

Recently the predominance of eosinophile cells has attracted some attention. Bine 2 
reported 2 cases of atypical pulmonary infection followed by pleural effusion. In 1 of 
these cases the blood contained 1% and the pleural effusion 51% of eosinophiles. Four 
years later, after the effusion had disappeared, the coarsely granular cells of the blood 
numbered 32%. In the second case these cells numbered 18% in the blood and from 
30% to 40% in the pleural effusion. His conclusion is that an eosinophilia of the pleural 
effusion precedes that of the blood, reaches a higher point than in the blood and is the 
first to disappear. The etiology of the cases with pleural eosinophilia is so varied that 
this formula can have little importance in diagnosis. The condition has been found 
following trauma and in typhoid fever, pneumonia, tuberculosis, syphilis, polyarthritis, 
general sepsis, pulmonary gangrene, endothelioma, hemorrhagic infarct of the lung, 
cardiac insufficiency, influenza, etc. A pleural eosinophilia and basophilia not infre- 
quently develop in cases of traumatic hemothorax. In some cases the eosinophilies 
number from 60% to 75% of the total cell count. They seem to be formed locally in 
the pleura and to be the result of a special transformation of leucocytic elements which 
can occur only in an aseptic medium. These cells later reach the blood and cause a 
tardy eosinophilia. It is believed that they are formed directly from the polynuclear 
finely granular leucocytes. 

Bacteriology. — The pathogenic micro-organisms in pleural exudates 
may, if present, be demonstrated in cultures or even in smears made directly 
from the fluid as it is removed. In cases following influenza we have seen a 
perfectly clear serous fluid from which grew Micrococcus aureus. The de- 
monstration of Bacillus tuberculosis, however, is exceedingly unsatisfactory 
because of the very small number of these organisms in the large volumes of 
the fluid. Jousett's method of inoscopy has been much used. At least 100 
c.c. of the fluid are allowed to clot, the clot enclosed in sterile gauze is then 
washed in sterile water and transferred to a flask. Then are added from 
ioto3oc.c. of a digestive fluid consisting of pepsin 1 to 2 gms., pure glycer- 
ine and concentrated hydrochloric acid, of each 10 c.c, sodium fluoride 3 
gms. and distilled water enough to make 1000 gms. This mass is digested 
in the thermostat at 3 8° C. until the clot has disappeared, which usually 
takes from 2 to 3 hours. This time may be shortened by occasionally 
shaking the mixture. The fluid is then decanted and smears are made 
from the sediment. Zebrowski recommends that 100 c.c. or more of the 
fluid be mixed with an equal amount of 1% sodium fluoride solution and 
kept in a cool place for 24 hours and then decanted. The sediment is 
then centrifugalized and smears made. Animal inoculation is, however, the 
only certain way of determining the presence of these bacilli. To be accu- 
rate, a large amount of the fluid must be injected into the peritoneal cavity 
of a guinea pig. According to some from 10 to 50 c.c. are injected each 
week until at least 300 have been introduced. The amount possible to 
inject will depend somewhat on its toxicity. The animal is then allowed 
to live for 3 months (unless it dies before then, in which case of course 
it is immediately autopsied, see page 27. At the end of this time the 
animal is killed and examined. Using this method Bacillus tuberculosis 

2 Am. J. of Med. Sc, 19 18, vol. 155, p. 579. 



EXAMINATION OF VARIOUS FLUIDS 703 

has been demonstrated in 85% of the cases of pleurisy with effusion 
(Le Damary said in 03%). 

Empyema. — In pneumococcus empyema which is primary or a sequela 
of lobar pneumonia the pus is creamy and sometimes so thick that none is 
obtained through a fine hypodermic needle. It is very rich in pus cells and 
usually contains many encapsulated diplococci. The pneumococcus 
found is usually type 1, 2 or 4. The empyema which develops during an 
attack of bronchopneumonia is an early rather than a late complication and 
so is often coexistent with the pneumonia, a matter of great moment in 
therapy since the evacuation of the fluid is a much more serious matter. 

Streptococcus hemolyticus empyema is the form of empyema which 
predominated in the army during the epidemic of 1 918-19. It is a rare type 
in civil life. The fluid on aspiration has a light yellow color with a greenish 
tinge and at first is only slightly cloudy and slightly purulent. The cocci 
are numerous, large, round or slightly flattened and in chains of from 2 to 
10. The cultures from the fluid on blood-agar plates show after 24 hours of 
incubation small, elevated, opaque colonies of a pearl-gray color, each sur- 
rounded by a sharply defined zone of hemolysis. Subcultures in blood 
broth give a diffuse and cloudy growth with a heavy sediment. The blood 
cells are rapidly destroyed therefore the broth becomes of a claret-red color. 
The smears of these cultures will show chains of streptococci. 

In Streptococcus viridans empyema the fluid is thin, grayish-red in 
color, turbid because* of fine fibrin flakes and much later becomes truly 
purulent. These non-hemolyzing streptococci were found both in the spu- 
tum and in the pleural exudate of these patients. 

In the influenza empyemas of 1917-20 the pneumococcus seemed to be 
the predominating organism and yet the pneumonia is very different from 
the ordinary lobar pneumonia since it was a very virulent necrosing 
bronchopneumonia. The influenza empyema developed late in the course of 
the pneumonia (in from the first [rarely] to the fourth week). These 
cases are frequently complicated by multiple metastatic abscesses in- 
various parts of the body. The exudate is small in amount and when, 
abundant enough to aspirate, often is too thick to flow through an 
ordinary trocar. 

Streptococcus empyemas sometimes complicate pneumococcus pneu- 
monias. Of the 80 cases of Brooks and Cecil 9 showed this mixed infection, 
while in 1 case the emypema was due to Staphylococcus aureus. Strep- 
tococcus hemolyticus and Streptococcus viridans were found in these fluids, 
while the lobar pneumonia was due to the second , third and fourth type of 
the pneumococcus. 

Sterile empyemas. — In about 20% of empyemas the fluid is sterile. 
In a majority of these cases it was purulent or semipurulent in character. 
Although the cultures are sterile, yet, in practically all cases disintegrated 
Gram-positive cocci can be seen in smears of the fluid. Most of these cases- 



704 CLINICAL DIAGNOSIS 

accompany a pneumococcus lobar pneumonia, the others, a strepto- 
coccus pneumonia. 

In hemorrhagic pleurisy the exudate contains so much blood that it is 
grossly apparent in the fluid. Such fluids are met with in cases of malignant 
disease involving the pleural surfaces; in tuberculosis, in which case a blood 
vessel is supposed to be eroded by the tuberculous process; in various as- 
thenic conditions including B right's disease; in cancer in other parts of the 
body; in cirrhosis of the liver; and in rare cases of the malignant type of 
acute infectious fevers. In some cases of influenza a bloody pleurisy has 
appeared on from the fifth to eighth day of the influenza. The pleural ef- 
fusion is almost pure blood. This character of the fluid seemed to have very 
little effect on the course of the disease. Bloody pleural effusions have been 
found also in healthy persons due to causes as yet undetermined. 

Trauma of a blood-vessel by the needle used in aspiration is a common 
cause of the bloody effusion. 

The pleural fluid of cases with carcinoma of the pleura is serous at first 
and possibly would remain so if not tapped, but later becomes blood- 
stained or hemorrhagic or stained by old blood and therefore of chocolate 
color. The amount of blood present would seem to depend in part on the 
degree of negative pressure used in the aspiration. 

In cases of primary sarcoma of the pleura there is usually an effusion 
present, hemorrhagic as a rule, or will be so at least after the chest has 
been once tapped. On microscopic examination of the sediment the 
discovery of spindle cells may suggest the diagnosis. This disease 
is uncommon. 

The majority of malignant tumors of the pleura are secondary to those 
of the lung, mediastinum or chest wall, while others are metastases from any 
distant organ. These tumors are as a rule, small, separate nodules. The 
pleural fluid in such cases may be serous but is generally bloody. 

In actinomycetic infection of the pleura this cavity usually contains 
pus in which can be seen grossly the sulphur granules (see page 45). Some 
of these are very minute in size, others even 2 mm. in diameter. They are 
white or yellow in color (hence the name "sulphur granule") and are irreg- 
ular in shape and soft, but some are calcareous in texture. 

Pericardial Fluid. — This fluid normally is of a lemon-yellow color, is 
slightly viscid and seems to contain more fibrin than do other physiological 
fluids. The solids it contains vary from 37.5 to 44.9 gms. per liter; the al- 
bumin, 22.8 to 24.7 gms.; soluble salts, from 8 to 9 gms., the insoluble 
salts, 0.15 gm. per liter, and extractives 2 gms. per liter. 

The fluid of a recent tuberculous case had a specific gravity of 1.0204; solids 5.8%; 
total proteid 3.91% and globulin only 15.2% of this. 

Synovial Membrane. — The synovial fluid is alkaline, thick, sticky, 
viscid , yellowish in color, and cloudy often from cell detritus ; or it is clear. 



EXAMINATION OF VARIOUS FLUIDS 705 

It contains albumin, salts and a body which is physically like mucin, but 
which it cannot be, since no reducing body can be split off. Neither is it 
nucleo-albumin. . Salkowski has given it the name "synovin." 

The fluid from a recent case of rheumatism, and which clotted firmly, had a total 
proteid content of 4.3%; water-soluble extractives 1.07% (ash. 0.606%); alcohol-ether- 
soluble extractives 0.076% (ash, 0.046%) and fat fraction 0.35%. 

Chylous and Chyliform Effusions. — Fluids which are milky in appear- 
ance because of a high fat content are called "chylous" fluids. These are 
evidently due to lesions of the thoracic duct or of its radials. Those whose 
milky appearance is due to large numbers of cells undergoing fatty de- 
generation are termed "chyliform". This distinction has, however, proven 
of little practical value, for the difference between these 2 kinds of fluids 
would seem to be of one degree rather than of kind. These effusions are 
rather rare. They have, been found in patients of all ages and in 1 or several 
serous sacs. These fluids are milky-red (from the mixture of blood), 
yellow or green in appearance and odorless or with a slightly sweetish odor. 
They may in appearance resemble pus. If allowed to stand a definite cream 
may rise. The emulsion of the fat cannot be cleared by filtration or by 
centrifugalization, but will disappear if the fluid be shaken out with ether. 
Chemical analyses have shown the following: 

Fat 0.06 to 3.71% 

Albumin 3.36 to 7.37% 

Total solids 5 to 10% 

Organic matter 1 % 

Casein and fibrin have been found as well as cholestrin and lecithin. 
Glucose may be present in about the frequency and amounts that it is 
present in clear serous effusions. If more than 0.2% is present, however, 
this may indicate the thoracic duct as the source of the effusion. Micro- 
scopic examination shows the fat in fine emulsion and (in the chyliform 
cases especially) degenerating cells in great numbers. 

Pseudochylous fluids are practically fat-free and owe their milky 
appearance to some unknown albuminous body. If allowed to stand these 
fluids remain homogeneously milky. In some cases the milkiness is due to 
albumin of the globulin group from which large amounts of lecithin can be 
extracted by hot alcohol. What fat is present resembles that of the so- 
called fatty degeneration of the epithelial cells. Quincke first showed that 
albumin in fine granules could give a fluid a milky appearance. 

Some fluids become more milky on cooling. In some cases a perfectly 
clear fluid on the first tapping becomes progressively more milky on the 
subsequent tappings. 

The specific gravity of these fluids varies from 1. 010 to 1. 014. Ini case 
it was 1 .06 1 and in another it was 1 .08 1 . In these cases much pus must have 
45 



706 CLINICAL DIAGNOSIS 

been present. Their reaction is alkaline and, strange to say, they have no 
odor. In the cases that we have examined this has been a marked feature. 
They are very resistant against decomposition and can remain in the lab- 
oratory for weeks without apparent change. The sediment is scanty, 
consisting of epithelial cells, all degenerated and full of fatty globules and 
globules which do not take the stains of fat. 

Seme of these cases of chylous effusion are due to filaria disease (see page 
673) while in others the fat may best be explained by the fatty globules freed 
from the fatty degenerated epithelium cells. But the majority of cases are 
hard to explain. Our best case "was one of tuberculosis of the peritoneum. 
Naunyn taught that amyloid degeneration of the vessels of the peritoneum 
was an important cause. In Tabora's case of peritonitis carcinomatosa the 
fluid contained 1.2% of fat and 0.864% of sugar. The opalescent fluids occur 
in a great variety of conditions and are often found at autopsy; in cachexias, 
heart cases, etc. The reason suggested for chylous fluids in cases of heart- 
failure is stasis in the thoracic duct. In other cases the stasis may be due to 
the pressure of tumors on the duct. 

In one case ot markedly chylous ascites we studied the specific gravity of the fluid 
was 1. 013 and the proteid 5. 114 gms. per liter of which 73% was globulin. The fat- 
cholesterin-lecithin-fraction was 1.469%. 

A chyliform ascitic fluid from a case of uremia had a specific gravity of 1.0055 an( l 
contained 1.2988% of solids. 

OVARIAN CYSTS 

Colloid as applied to the contents of ovarian cysts is not the name of one 
substance but of a group of fluids with certain physical properties, which 
is found in various cysts and organs. These fluids are gelatinous, the 
gelatinous substance insoluble in water and acetic acid but soluble in 
alkali. From some may be split off a reducing body but their composition 
varies much. 

Pseudomucin (Metalbumin). — Pseudomucin occurs in many of the 
ovarian cyst contents which are very viscid and slimy. Alcohol gives a 
thready precipitate resembling wood-pulp, which can be wound around the 
rod. It is not precipitated by heat nor by acetic acid. The precipitate 
formed by alcohol is ground fine under alcohol and then freed from alcohol 
by means of ether, and dissolved in water. It is then reprecipitated with 
alcohol. A light white powder is obtained which is soluble in water to an 
opalescent mucoid solution, which is not well precipitated by acetic acid. 
When boiled with HC1 an abundant reducing body is split off which reduces 
copper very easily. 

Paramucin. — Paramucin is a substance present in certain ovarian cysts, 
also in the ascitic fluid providing the ovarian cyst has already ruptured into 
the abdomen. It is firm, glistening, with the consistency of gelatin, soluble 
in dilute m'neral acid, shrinks in acidulated alcohol and in alcohol and ether, 



EXAMINATION OF VARIOUS FLUIDS 707 

and can be reduced to a fine white powder. Its characteristics are: its 
insolubility in water, the fact that it swells in alkali dissolving in excess, that 
it is precipitated by acetic acid and is soluble in excess and, especially, that 
it will reduce copper salts without preliminary boiling with acid. 

Serous Cysts (dilatation of the Graafian follicles) contain a perfectly 
clear serous fluid, watery, which foams easily, has an amber color and a 
specific gravity of from 1.005 to 1.022 (usually 1.005 to 1.014). It contains 
from 10 to 40 gms. of solids per liter and all the various constituents 
of serous fluids. 

In 2 recent cases the specific gravity was 1.022 in 1 case and 1.016 in the other. 
They contained a great deal of albumin. Heat alone caused but a faint cloud, but I 
drop of acid made the fluid perfectly solid. Both serum globulin and serum albumin 
were present, but little or no euglobulin. 

Proliferating Cysts from Pfliiger's Tubules. — The contents of the cysts 
from Pfliiger's tubules vary much. Some contain "colloid," which on 
boiling with acid gives a reducing body. From the colloid the fattycrystals 
(soluble on warming) may be watched as they crystallize out singly and 
in rosettes (see Fig. 149). 

Another group of these cysts contain a viscid fluid, very stringy, which 
varies much in consistency. It is of a brownish or dark greenish-brown 
color. 

The specific gravity of the contents of 2 cases was 1.025 an( l 1-0302; the solids 
were 9.7 and 9.3% and the alcohol precipitate, 6.9 and 8.5%. 

Some, however, contain a thin watery fluid, of a bluish- white opalescent 
color which may, however, be yellowish, yellowish-brown, or greenish, 
according to the amount of blood present. 

We give a few examples of the contents of such cysts which we have seen. 

1. The fluid was quite opalescent; specific gravity, 1.0043; solids 2.837%. The 
alcohol precipitate was 1.98% of the fluid weight, resembled macerated filter paper, 
was not stringy, could be reduced to a fine white powder and was with difficulty sol- 
uble in water to an opalescent fluid. Watery extractives 0.524 (ash. 0.388)%, alcohol- 
ether soluble extractives 0.2056 (ash. 0.108)%; fat, cholesterol etc., 0.96%. 

2. This fluid was reddish-yellowish in color and contained considerable sediment 
of small epithelial and some large epithelial cells with coarse refractile granules. It 
filtered clear. Its specific gravity was 1.0080, the solids 2.32% and the alcohol precipi- 
tate similar to the above. 

3. This fluid had a bluish opalescent appearance and a specific gravity of 1.0073. 

The contents of some of the multilocular cysts are thick, of a yellowish- 
red or brown color depending on the blood pigment, not specially viscid and 
contain a suspension of glistening masses of cholesterol crystals (see Fig. 150). 

In a recent case the figures were: specific gravity 1.0259 and the alcohol precipitate, 
10.56%; it contained no reducing body. Half saturation of the original filtered fluid 
with (NH 4 ) 2 S0 4 gave a precipitate of 0.692%, (globulin?): albumin (?) 0.604%, extrac- 



708 CLINICAL DIAGNOSIS 

tives, soluble in water, 1.46% (ash. 0.266%); alcohol-soluble extractives, 0.56% (ash. 
0.44%). Microscopically there was a great amount of detritus in the sediment mixed 
with very large cells (epithelial) full of glistening granules, cholesterol crystals and 
fat needles. 

In another similar case the specific gravity of the fluid was 1 .0306. 

The sediment contains much detritus, red blood-cells, leucocytes, large epithelial 
cells, single and in groups and filled with granules like fat, large masses of fatty granules, 
cholesterol crystals and colloid granules which are large, circular, strongly 
refractive bodies. 

One DERMOID Cyst contained much paramucin, also serum globulin and 
albumin. The water-content of the jelly of this case was 92.2%, its alcohol precipitate 
3.3%, the water-soluble extractives, 0.4% (ash. 0.27%) and the alcohol-ether extract- 
ives 0.25% (ash. 0.16%). 

Tubo-ovarian Cysts. — The contents of the tubo-ovarian cysts is a 
watery, thin, serous fluid which contains no pseudomucin. 

Parovarian Cysts. — The parovarian cysts contain a thin, watery fluid 
which is colorless or has a very pale yellow color or is slightly opalescent. 
Its specific gravity varies from 1.002 to 1.009, "the solids from 10 to 20 gms. 
per liter. It contains no pseudomucin. Albumin may be entirely 
absent or be present in traces only. These contain therefore only 
water and ex-tractives. 

In a recent case, that of a patient aged 20 years, the cyst contained about 2 liters 
of very clear watery fluid with very slight opalescence. Its specific gravity was 1 .0078, 
only the faintest precipitate with alcohol or ammonium sulphate could be obtained 
and the chlorides were 0.45% (as NaCl). Microscopically it contained very few epithe- 
lial cells, which were round and granular and with a round nucleus. 

Intraligamentous Cysts. — The fluid of the intraligamentous cysts 
is yellow, yellowish-green, or brownish in color, contain little or no 
pseudomucin, its specific gravity varies from 1.032 to 1.036, its solids 
from 90 to 100 gms. per liter and the proteids it contains are those of the 
blood plasma. 

HYDROCELE 

The fluid of hydroceles is clear, dark yellow or greenish in color, the 
specific gravity varies from 1.014 to 1.026 and the solids average 60 gms. 
per liter. This fluid sometimes coagulates spontaneously. Leucocytes are 
always present in it and sometimes cholesterin crystals. 

For illustration, in one case the specific gravity of the contents of the hydrocele was 
1. 0107; the solids, 6.329%; total albumin, 5.29% of which 45% was globulin; the water- 
soluble extractives were 0.7504% (ash. 0.462%); the alcohol-ether extractives, 0.452% 
(ash. 0.1726%); and the fat fraction 0.1864%. 

SPERMATOCELE 

The fluid of spermatocele cysts is colorless, watery and slightly milky. 
Its specific gravity is from 1.006 to 1.010. The solids average 13 gms. per 



EXAMINATION OF VARIOUS FLUIDS 709 

liter, the proteids are slight in amount, while the sediment contains cells 
detritus, fat granules and spermatozoa. 

The TOPHI of gout, so important in diagnosis, can be distinguished from 
small sebaceous cysts, small cartilaginous tumors, etc., only by the micro- 
scopic examination of their contents. A little of this mixed with water is 
found to be an amorphous paste containing many needles of sodium bi- 
urate (Fig. 151). The sebaceous cysts on the other hand contain amor- 
phous sebaceous matter, containing many fatty and cholesterin crystals. 

The masses of urea crystals, the " urea frost," which appear on the 
skin of the face just before death in rare cases of nephritis (there have been 
but 5 cases in this clinic) may be tested by the methods given on page in. 
This is a most interesting phenomenon. The circulation in the skin is so 
poor when it occurs that it is very hard to believe that the immediate 
source of this urea can be the blood. 



INDEX 



Abscess, leucocytes in, 516 
of liver, blood in, 656 

sputum in, 79 
of lung, leucocytes in, 516 
sputum in, 78 
Absolute amount of HC1 in 

gastric juice, 349 
Accidental albuminuria, 227 
Acetic acid in gastric iuice, 

359 
Aceto-acetic acid, 192 
Acetone, quantitative deter- 
mination of, 189 
in the urine, 187 
Acetone bodies, determination 

of, 197 
Acholic stools, 389 
Achromatophilia, 475 
Achylia gastrica, 372 
Acid, alloxyproteinic, 128 
chrysophanic, 99, 153 
diacetic, in urine, 192 
glycuronic, 162, 206 
hippuric, 253 
homogentisinic, 98, 208 
hydrochloric, 92 
lactic, 357, 377, 695 
nitric, 140 
nitrous, 140 
oxalic, in urine, 250 
oxybutyric, 194 
oxyproteinic, 128 
phosphoric, 133 
silicic, 140 
sulphocyanic, 140 
sulphuric, 136 
thisulphuric, 140 
uric, 114 
uroleucinic, 208 
Acid-fast bacteria, 29 
Acidity of gastric juice, 344; 
determination of 349; of 
urine, 100; determination 

of, 103 
organic, in gastric juice, 

359 
percent 346 

total, of gastric juice, 
345 
Acidophilic cells, 475 

leucocytes, 492 
Acidosis of diabetes, 202 
Acids, in urine, 155 

fatty crystals, in sputum, 
18 
Acromegaly, blood in, 647 
Actinomycosis of lung, sputum 

in, 45; of pleura, 704 
Acute articular rheumatism, 
blood in, 644 
bronchitis, 68 

diffuse nephritis, of 
cholera, 329 
diffuse nephritis, of lues, 

urine, in, 329 
diseases, blood in, 630 
gastritis, 369 
leukemia, 625 
lobar pneumonia, sputum 

of, 90 
luetic nephritis, urine in, 
329 

miliary tuberculosis, 

blood in, 637 
sputum in, 55 



Acute nephritis, urine in, 327 
nephritis of cholera, urine 

in, 329 
parenchymatous nephri- 
tis, 329 
pneumonic tuberculosis, 

55 
purulent gastritis, 370 
rheumatic fever blood in, 

644 
yellow atrophy of liver, 
blood in, 657 
Addison's disease, blood in, 

638, 658 
Adenin, 120 
Adenitis, tuberculosis, blood 

in, 628 
Adolescence, albuminuria of, 

227 
Adrenals, tuberculosis of, 

blood in, 638 
^stivo-autumnal malaria par- 
asite of, 664 
Agar media, 285 
Agchylostoma duodenale, 409 
Age, effect of, on count of reds, 

483 
Agglutination phenomena, 564 
Agglutinins, 564; blood, 588 
Agonal leucocytosis, 518 
Air, bad, anemia due to, 597 

in sputum, 5 
Albumin, calculi of, 2 79 

determination of, in body 

fluids, 693 
in gastric contents, 356 
in stools, 396 
quotient, 217 
serum, 218 
tests in urine, 210 
to remove from urine, 217 
Albuminous expectoration, 81 
Albuminuria, 217, 224 325 
accidental, 227 
adolescence, 227 
after baths, 233 
alimentary, 226 
anaemia due to, 599 
cyclic, 227, 228 
due to Bright's disease, 

232 
due to definite renal 

lesion, 232 
due to palpation of kid- 
ney, 231 
essential, 227 
false, 225 
febrile, 231 
functional, 225 
haematogenous, 231 
hereditary, 230 
hypostatic, 230 
intermittent, 230 
luetic, 231 
minima, 230 
nervous, 231 

of apparently healthy, 227 
of diabetes, 205 
of labor, 227 
of masturbators, 227 
of new-born, 227 
of pregnancy, 22 7 
of puberty, 227 
orthostatic, 227 
orthotic, 227 
physiological, 224 
post-infectious, 230 
postural, 227 



Albuminuria, splenic, 228 

structural, 225 

thermolytic, 233 

traumatic, 231 

true, 224 

without definite renal les- 
ion, 224 
Albuminuric cicatricielle, 230 

paracellaire, 230 

phosphaturique, 230 

pregoutteuse, 230 

residuale, 230 
Albumosuria, 235 

alimentary, 236 

enterogenous, 236 

febrile, 236 

haematogenous, 236 

hepatogenous, 236 

myelopathic, 233 

pyogenic, 236 
Aleppo button, 673 
Alimentary albuminuria, 226 

albumosuria, 2Zf> 

glycosuria, 162 

levulosuria, 182 

lipuria, 262 
Alkaline tide, 101 
Alkalinity of blood, 530 
Alkaptonuria, 207 
Allantoin, 698 
Alloxuric bases, 120 
Alloxyproteinic acid, 128 
Almen's test for glucose, 170 
Aloin test, 394 
Altitude, effect of, on count of 

reds, 485 
Alveolar epithelial cells in spu- 
tum, 12, 68 
Alymphemic lymphomatosis, 

521 
Amboceptors, 590 
Ammonia, 122, determination 
of, 123 

in gastric contents, 359 

in urine, 122 

of blood, 542 
Ammonium biurate, 245 

magnesium phosphate, 248 
Amoeba coli, 401, 402 

dysenteriae, 401 

mitis, 403 

pulmonalis, 50 

vulgaris, 403 
Amoebic dysentery, 401 
stools in, 423 

anasmia due to, 598 

eosinophilia due to, 523 

gastric contents in, 351 
Amount, absolute, of HC1 in 
gastric juice, 349 

of sputum, 2 

of urine, 89 
Amyloid kidney, urine in, 333 
Alkali reserve of plasma, 534 

starvation, 202 
Alkaligenes bacillus, 288 
Alkalinuria, 101 
Allen's paradoxical law, 164 
American hook worm, 410 
Ammoniacal silver magnesium 

mixture, 119 
Amphophilic granules, 492 
Amylase in urine, 160 
Anasmia, 590 

aplastic, 592, 615 

chlorotic,_ 592 

consumptive, 592 

due to acute gastritis, 598- 

711 



712 



INDEX 



Anaemia due to acute hemor- 
rhage, 594 
due to acute infections, 

599 
due to albuminuria, 599 
due to amoebic dysentery 

598 
due to ascaris, 601 
due to bad air, 597 
due to blood poisons, 596 
due to Bothriocephalus 

latus, 600 
due to chronic gastritis, 

598 
due to chronic hemor- 
rhage, 595 
due to chronic infectious 

diseases, 599 
due to coal-tar products, 

602 
due to congenital lues, 

653 
due to constipation, 598 
due to cryptic septi- 
cemia, 599 
due to diarrhoea, 598 
due to gastro-intestinal 

disorders, 598 
due to hemorrhage, 594 
due to inanition, 596 
due to intestinal para- 
sites, 600 
due to lactorrhea, 599 
due to lack of sunlight, 

597 
due to latent infection, 

598 
due to lead, 601 
due to malaria, 559 
due to marasmus, 599 
due to overwork, 597 
due to poisons, 601 
due to poor food, 597 
due to pus formation, 599 
due to rickets, 647 
due to spermatorrhea, 599 
due to starvation, 597 
due to Strongyloides in- 

testinalis, 600 
due to tape worm, 601 
due to ulcerative colitis, 
598 

due to uncinariasis, 600 
due to worry, 597 
due to yeasts, 601 
due to yellow fever, 599 
hemolytic, 592 
hypoplastic, 592 
of children, 645 
of growth, 645 
of inanition, 596 
of the poor, 596 
of the South, 410 
of the Tropics, 598 
posthemorrhagic anaemia, 

594 
primary, 591 

primary pernicious. 602 
Pseudoleukaemica infan- 
tum, 646 
secondary, 592, 598 
simple primary, 602 
splenic, 602 
Anaemic degeneration of cells, 

475 
Anaglycosuria, 324 
Analine gentian violet, 35, 38, 

281 
Ancestral corpuscles, 505 
Anchovy sauce sputum, 7 9 
Anchylostoma duodenale, 410 
Angina of Plant, 87 

Vincent, 87 
Angina ulceromembranous, 87 
Angioneurotic haematuria, 237 
Anguillula aceti in urine, 307 
intestinalis, 413 
stercoralis, 413 



Animal gum, 162, 187 

parasites in sputum, 50 
in stools, 401 
in urine, 306 
Ankylostomum duodenale, 409 
Anopheles mosquito, 667* 
Anthracosis, 4 
Anthrax leucocytosis in, 514 
Antiformin method, 24 
Antigens, 576 
Anuria, 90, 92 
Appearance, general, of urine, 

88 
Appendicitis, blood in, 644 
Aplastic anemia, 615 
Arabinose in urine, 184 
Ameth's classification of leu- 
cocytes, 508 
Arsenic in urine, 143 
Arteriosclerosis of kidney, al- 
buminuria in, 325 
urine in, 332 
Arthritis deformans, blood in, 

645 
Arthus and Huber method 

(trypsin), 382 
Ascaris lumbricoides, 53, 407 

anemia due to, 601 
Ascitic fluid, cyto-diagnosis of, 

700 
Asiatic cholera, stools in, 422 
Aspergillus flavus, 47 

fumigatus in sputum, 47 
glaucus, 48 
niger, 48 
subfuscus, 48 
Aspirated foreign bodies, 78 
Aspiration pneumonia spu- 
tum of, 65 
Assimilation limit for fat, 425; 

for sugars, 162 
Asthma, blood in, 643 
eosinophilia in, 522 
humidum, 72 
sputum of, 66 
Atony of stomach, 364 
Atrophy, acute yellow, of liver, 
blood in, 657 
of mucosa, 372 
renal, 331, 335 
senile of kidney, 335 
Auer's bodies, 628 
Avian tuberculosis, 29 
Avirulent diptheria bacillus, 

86 
Azotorrhoea, 396, 425 



B 



Bacillary dysentery, 401, 424 
Bacillus aerogenes capsulatus, 

289 
alkaligenes, 288 
bifidus, 419 
Bordet's, 37 
buccalis maximus, 32 
coli communis, 286 
diptheriae, 37 

avirulent, 40 
dysenteries, 424 
fusiformis, 41 
influenzae, 34 
Kauffman's, in gastric 

juice, 379 
lactis aerogenes, 288 
leprae, 29, 31 
mucosus capsulatus, 34 
paratyphosus, 287 
proteus, 288 
pseudodiphtheriae, 40 
pyocyaneus, 5, 289 
tetani, 290 
tuberculosis, 22 

bovine type, 28 

in sputum, 22 

in stools, 420 

in urine, 283 
typhosus, 287, 421 



Bacillus ulceris cancroci, 300 
virescens, 5 
Y, 424 
Bacilluria, 291 
Bacillus-carriers, 291 
Bacteria, acid fast, 29 

acidophilic, 419 . 

in gastric contents, 362 

in sputum, 18 

in stools, 387, 419 

in urine, 279 

media for, 35 

stains for, 280 

thermophilic, 419 
Bacterial casts, 268 
Bacteriology, of blood, 561 

of pleural fluid, 703 

of sputum, 18 

of urine, 279, 290 
Bacteriorrhcea, 297 
Bacteriuria, "294 
Bacterium termo, 289 
Bacteriuria, 294 
Balantidium coli, 406 

in urine, 306 
Bases, alloxuric, 120 

inorganic, in urine, 129 

nuclein, 120 

of gastric juice, 359 

purin, 120 

xanthin, 120 
Basophile granules in red cells, 
479 

of leucocytes, 492 
Basophilia, 475 
Basophilic red cells, 501 
Baths, albuminuria following, 
227 

effect of, on red count, 487 
Bence-Jones's body, 233, 253 
Benedict's method for deter- 
mination of glucose, 176 
Benzidin test for blood, 394 
Betaoxybutyric acid, 194 
Bial's test for pentose, 185 
Bile acids in urine, 155 

drainage, 383 

in urine, 148 

in vomitus, 350 

pigments in urine, 97, 148 

stained sputum, 4 

to remove from urine, 154 
Bilharzia eggs in urine, 306 
Bilifuscin, 149, 150 
Biliprasin, 149, 150 
Bilirubin, in blood, 529 

in stools, 289 

in urine, 148 

in urine sediment, 253 
Biliverdin, 149, 150 
Birds, tuberculosis of, 29 
Bismarck brown, 35 
Bismuth crystals in stools, 398 
Biurate of ammonia, 245 
Biurates, 116 
Biuret test, 111 
Black urines, 98 
Blackwater fever, 238 
Bladder stones, 276 

tuberculosis of, 292 
Blastomycetes, 42 

in blood, 601 

in stools, 418 

in urine, 305 
Blastomycosis, 42 
Bleeding occult, 394 
Bleeding time, 437 
Blood, 428 

agar, 285 

agglutination, 588 

alkalinity of, 530 

ammonia of, 542 

bacteriology of, 561 

bilirubin in, 529 

carbon dioxide combining 
power of, 532 
, cells, red, 445 
white, 450 



INDEX 



713 



Blood, chemical tests for, 240 
chlorides of, 538 
cholesterin of, 558 
coagulation of, 431 
creatin, 549 
creatinin, 547 
crises, 501 

diastatic activity, 554 
dried residue, 431 
dust, 451 
fat in, 452, 561 
fibrin, 452 
forceps, 428 

fresh, study of, 445, 450 
glucose in, 549 
groups, 588 
hemoglobin, 439 
hydrogen ion concen- 
tration of, 532 
in abscess of liver, 656 
in acromegaly, 647 
in acute articular rheuma- 
tism, 644 
in acute diseases, 629 
in acute yellow atrophy, 

657 
in Addison's disease, 658, 

639 
in appendicitis, 654 
in arthritis deformans, 

644 
in bronchial asthma, 643 
in bronchopneumonia, 643 
in cancer, 649 

of breast, 650 
of intestines, 652 
of oesophagus, 651 
of rectum, 652 
of stomach, 650 
of testicle, 652 
in carbon monoxide poi- 
soning, 530 
in catarrhal jaundice, 656 
in cholangitis, 656 
in cholecystitis, 656 
in chorea, 647 
in chronic diseases, 647 
in chronic septicaemia, 618 
in cirrhosis of liver, 657 
in depressive insanity, 647 
in diabetes mellitus, 647 
in diptheria, 635 
in diseases of liver, 656 
in endocarditis, 632 
in gall-stone colic, 656 
in gastric contents, 363 
in general paresis, 647 
in German measles, 633 
in heart disease, 657 
in inanition, 658 
in influenza, 643 
in leprosy, 658 
in liver abscess, 656 
in liver disease, 656 
in lues, 652 

in lues, congenital, 646 
in malaria, 630 
in malignant disease, 649 
in maniacal depressive in- 
sanity, 647 
in measles, 633 
in meningitis, 644 
in myxcedema, 658 
in nephritis, 654 
in nervous diseases, 647 
in pneumonia, 640 
in renal diseases, 654 

acute nephritis, 654 
bilateral cystic kid- 
ney, 656 
chronic nephritis, 655 
in rickets, 658 
in sarcoma, 652 
in scarlet fever, 633 
in scurvy, 658 
in septicaemia, 631 
in smallpox, 634 
in stools, 393 



Blood in summer diarrhoeas, 647 
in intestinal parasites, 643 
in tonsillitis, 635 
in toxic jaundice, 656 
in tuberculosis, 634 
acute miliary, 637 
of adrenals, 638 
of bones and joints, 
of intestines, 638 
of lungs, 635 
of lymph glands, 637 
of meninges, 637 
of peritoneum, 638 
of serous membranes, 
636 

renal, 638 
in typhoid fever, 638 
in typhus fever, 632 
in vomitus, 340 
lecithin in, 556 
leucocyte counting, 461 
leucocytes, 450 
lipoids of, 555 
Miiller's blood dust, 451 
nonproteid nitrogen, 540 
pigments, in, 439 
reaction of, 530 
red cells of, 445 
sedimentation, 431 
specific gravity of, 429 
test for, Almen's, 241 

guaiac, 241 
haemin, 240 
Heller's, 240 

Sehonbein's, 241 
spectroscopic, 241 
Teichmann's, 241 
urea in, 571 
uric acid in, 543 
urea in, 541 
urobilin in, 529 
viscosity of, 437 
Blood-casts, 267 
Blood-cells in gastric con- 
tents of cancer of stom- 
ach, 380 
in gastric juice, 363 
in sputum, 14 
Blood-cells in stools, 393 
in urine, 97 
in vomitus, 340 
Blood counting, 452 
Blood-crises, 501, 606 
Blood-cultures, 562 
Blood plasma, alkaline reserve, 

534 
Blood-platelets, 452 
Blood-serum, Loffler's, 37 
Blood-smears, 462 
Blood-staining, 466 
Bloody sputum, 3 
Blue, indigo, 145 

urines, 99 
Boas bacillus (Oppler), 362 

378 
Bodies, foreign, in sputum, 26 
Body fluids, examination of, 

693 
Bogg's coagulometer, 435 

methods for preserving 
and mounting worms, 
426 
throttle, 453 
Bone marrow, 500 

diseases of, eosino- 

philia in, 521 
white cells in, 504 
Bones, tuberculosis of, blood 

in, 638 
Bordet's bacillus, 37 
Bordet-Gengou phenomenon, 

569 
Bothriocephalus latus, 417 

cause of anaemia, 600 
Bottcher's crystals, 18 

medium, 37 
Bouillon, 286 

glycerine, 29 



Bound HC1, 344 

Bovine type, tubercle bacillus, 

28 
Bradshaw's albumosuria, 233 
Bradyuria, 206 

Brain tumors, spinal fluid, 689 
Brassy cells, 447 
Breakfast test, 342 
Breast, cancer of, blood in, 

650 
Bremer's blood test, 648 
Bricklayer's anemia, 410 
Brick-red sputum, 62 
Bright's disease, albuminuria 

of, 232 

blood in, 654 
Brodie-Russel coagulometer, 

435 
Bronchi, lues of, 74 
Bronchial asthma, blood in, 
643 

catarrh, desquamative, 69 

colic, 10 
Bronchiestasis, 74 

hemorrhage in, 76 

leucocytosis in, 516 
Bronchioliths, 10 
Bronchitis, acute, 68 

capillary, 70 

chronic, 70 

croupous, 73 

eosinophiles in, 11 

fetid, 73 

fibrinous, 73 

idiopathic fibrinous, 73 

leucocytosis in, 516 

plastic, 73 

putrid, 72 

with cardiac disease, 72 

with emphysema, 71 
Bronchoblennorrhcea, 72 
Broncho-pneumonia, blood in, 
643 

sputum of, 65 

tuberculous, sputum of, 
55 
Broncho-pneumonomycosis, 

45 
Bronchorrhcea, 72 

serosa, 72 
Buccalis maximus, bacillus, 32 
Buerger's capsule stain, 33 
Buerker's chamber, 455 
Butyric acid in gastric con- 
tents, 359 
Butyric oxy-acid in urine, 194 



Cabot's ring bodies, 477 
Cachexia, effect of, on red 
count, 487 

hypochlorite, 145 
Cadiverin, 257 

Calcified masses in sputum, 9 
Calcium carbonate concre- 
tions, 278: sediments, 
249 
of urine, 140 

oxalate concretions, 278 
crystals in sputum, 18 
in urine, 250 
phosphate sediments, 249 

stones, 278 
sulphate sediments, 252 
urate sediments, 276 
Calculosa, pseudophthisis, 10 
Calculus, renal, 339 _ 

leucocytes in, 516 
ureteral, 338 
California disease, 44 
Cancer fragments in gastric 
juice, 363 
in urine, 284 
leucocytes in, 517 
of bones, blood in, 650 
of breast, blood in, 650 
of intestines, blood in, 650 



714 



INDEX 



Cancer of kidney, urine in, 338 
of lung, sputum of, 83 
of oesophagus, blood in, 

652 
of rectum, blood in, 652 

stools in, 423 
of stomach, blood in, 363 
flagellates in, 380 
gastric juice in, 375 
of testicle, blood in, 652 
Capillary bronchitis, 70 
Capsule stains, 32 
Carbohydrate tolerance, 551 
Carbohydrates in stools, 396 
in urine, fermentable, 162 
unfermentable, 162 
Carbolfuchsin, 25, 41, 281 
Carbol-thionin stain, 469 
Carbon dioxide combining 

power of plasma, 532 
Carbon granules in sputum, 12 
Carbonates in sediments, 248 
in stools, 396 
in urine, 140, 162 
Carbon-monoxide poisoning, 
effect of, on red count, 529 
Carcinoma, blood in, 5 ±7 
Carcinoma fragments in urine, 

274 
Cardiac disease, blood in, 657 

gastric contents in, 351 
Carnin, 120 
Carriers, ^Bacilli, 291 
Cartilage fragments in spu- 
tum, 6 
Casts, 264 

bacterial, 268 
blood, 267 
chemistry of, 270 
colloid, 267 
combined, 268 
diagnostic importance of, 

270 
epithelial, 264 
fatty, 266 
fibrin, 266 

fibrinous, in sputum, 9 
glassy, 267 
granular, 265 
hemoglobin, 265, 267 
hyaline, 267 
of moulds in sputum, 45 
origin of, 268 
prostatic, 273 
pseudocasts, 268 
pus, 267 
size of, 264 
staining, 272 
testicular, 273 
urate, 268 
waxy, 266 
Catarrh, desquamative, of 
bowel, 397 
desquamatory, bronchial, 

69 
dry, 71 
sec, 71 
Catarrhal jaundice, blood in, 
656 
gastric contents in, 357 
pneumonia, desquama- 
tory, 13 
Cavity formation in tubercu- 
losis, 5 7 
Centrifugalization of urine, 

280 
Centrifuge, quantitative deter- 
mination of albumin, 216 
Cercomonads in sputum, 51 
in stools, 406 
in urine, 306 
Cercomonas coli, 405 
hominis, 404, 406 
intestinalis, 405 
see spinal fluid 
Cerebral hemorrhage spinal 
fluid in, 689 



Cerebral vomiting, 339 

change of reaction test, 
29 
Cerebrospinal fluid, 678 

meningitis, leucocytes in, 
513; spinal fluid in, 689 
Cestodes, 415 
Chalicosis, 5 

Chancres, organisms of, 300 
Character of sputum, 3 
Charcot-Leyden crystals in 
blood, 620 
in sputum, 18, 67, 74 
in stools, 398 
Chemistry of blood, 528 
of casts, 2 70 
of sputum, 53 
Children anemia of, 645 
blood of, 646 
leucocytosis of, 520 
leukemia of, 628 
malaria of, 646 
sputum of, 2 
Chloride, excretion in urine, 
129 
retention, 129 
Chlorides of blood, 528 
Chlorine crises, 129 
Chloroma, 4 
Chlorosis, 612 

Egyptian, 410 
Chlorotic, anaemia, 592 

cells, 475 
Chloruremia, 130 
Cholangitis, blood in, 656 
Cholecyanin, 149, 150 

test for bile, Stokvis, 154 
Cholecystitis, blood in, 656 

gastric juice in, 351 
Cholelithiasis gastric contents 

in, 351 
Cholera, acute nephritis of, 
329 
leucocytes in, 515 
stools in, 422 
Cholera infantum, blood inj 

647 
Cholera spirillum, 422 

Asiatic, leucocytosis in, 
514 
Cholesterol in sputum, 18 
in body fluids, 694 
in stools, 398 
in urine sediment, 253 
Cholesterinuria, 253 
Choletelin, 149, 150, 556, 558 
Cholin, 683 
Chondroitin sulphuric acid, 

162, 222 
Chorea, blood in, 633 

spinal fluid in, 688 
Chromogenic bacteria in spu- 
tum, 5 
Chromogens, color due to, 5, 
19 
in urine, 94 
Chronic bronchitis, 70 

diseases, blood in, 647 
gastritis, 370 

idiopathic fibrinous bron- 
chitis, 73 
interstitial pneumonia, 

sputum of, 65 
nephritis, 656 
passive congestion of lung 
sputum of, 83 
urine in, 327 
ulcerative tuberculosis, 
sputum in, 56 
Chrysophanic acid in urine 

99, 153 
Chyliform fluids, 705 
Chylous fluids, 705 
Chyluria, 99, 209, 259 
Cicatricielle albuminuric, 230 
Cinaenomonas hominis , 406 
Cipollina's test, 172 
Cirrhosis of liver, blood in, 657 



Clap threads, 273, 296 
Clay-colored stools, 389 
Cloudy swelling of kidney,. 

urine in, 326 
Coagula of albumin in stools, 
396 
of fibrin in sputum, 63 
Coagulation of blood,- 431 

time, 433 
Coal pigment in sputum, 4 
Coal-tar products, anaemia due 

to, 602 
Coarsely granular cells, 451 
Coccidioidal granuloma, 44 
Coccidiosis, 44 
Coccidioxes-immitis, 44 
Coccus radiaire, 360 
Coefficient of Ambard, 316 

of Haser, 94 
Coffin lid crystals, 248 
Colic, bronchial, 10 
Colitis, anaemia due to, 582 
gastric contents in, 351 
Collection of urine, 88 
Colloid, 707 

casts, 267 
Colloidal gold test, 685 
Colon bacillus, 286 
Color index of blood, 490, 59 l r 
608 
of red cells, 409, 591 
of sputum, 3 
of stools, 389 
of urine, 94 

due to medicines, 99 
Coma, diabetic, 202 

leucocytes in, 518 
Combined casts, 268 
Comma bacillus, 422 
Complement, fixation of, 569 
for gonorrhea, 584 
for lues, 569 
for tuberculosis, 586 
Concretions, 276 

acid calcium phosphate, 

278 
albumin, 2 79 
ammonium urate, 245 
bladder, 2 76 
calcium carbonate, 278 

oxalate, 278 
cystin, 278 
fatty, 279 
in bowel, 398 
indigo, 279 
phosphate, 2 78 
renal, 276 

table for detection of, 277 
triple phosphate, 248 
uric acid, 276 
vesical, 276 
xanthin, 278 
Complement, 5 72 
Conductivity, electrical, 311 
Confinement, albuminuria of, 

227 
Congenital cystic kidney, 335 
heart disease, effect of, 

on red count, 657 
lues, blood in, 646 
Congestion, chronic, passive, 

of lung, 83 
Congo red paper, 345 
Conradi (Drigalski and) med- 
ium, 421 
Consistency of sputum, 3 

of stools, 387 
Constipation, 388 

anaemia due to, 598 
gastric contents in, 351 
latent, 387 
Constituents of normal stools, 

387 
Consumptive anaemia, 591 
Continous secretion, 367 
Contracted kidney, 331 
Cord, compression of, 689 
Cornil's marrow cells, 504 



INDEX 



715 



Cornil's myelocytes, 619 
Corpora amylacea in prostatic 
fluid, 308 
oryzoid.es, 57 
Cough, whooping, sputum of, 

66 
Counting-chamber, 455 
Counting leucocytes, 461; red 

cells, 452 
Cover glasses, 428 
Craw Craw, 675 
Creatin, 128 

in blood, 548 
Creatinin, 125 

in blood, 547 
Crenated red cells, 446 
Crescents, 665 
Crescentric bodies in red cells, 

449 
Cresyl blue, 478 
Crises, blood, 501, 606 
Cross fixation, 587 
Crotonic acid, 195 
Croupous bronchitis, sputum 
in, 70. 
pneumonia, blood in, 640 
sputum of, 60 
Cryoscopy, 310 
Crystals, Charcot-Leyden, in 
blood, 619 
in sputum, 18 
in stools, 398 
fatty acid, in sputum, 18 
in gastric juice, 363 
in sputum, 18 
in stools, 397 
melting point, determin- 
ation of, 173 
spermin, in prostatic fluid, 
309 
Culex mosquitoes, 664 
Cultures of blood, 562 
of sputum, 18 
of threat, 85 
of Treponema pallidum, 

301 
urine, 284 
Curds in stools, 396 
Curschmann's spirals, 7, 67 
Cyanosis, effect of, on red 

count, 487 
Cyclic; vomiting, 339; albumi- 
nuria, 227, 228 
Cylindrical epithelial cells in 
sputum, 12 
epithelium in stools, 414 
in urine, 262 
Cylindroids, 267 
Cylindruria, 271 
Cystic kidney, congenital, 
urine in, 335 
blood in, 656 
Cystin in urine, 256 

stones, 278 
Cystinuria, 256 
Cystitis, 291 

proteus, 293 
streptococcus, 293 
tuberculous, 291 
Cysts, echinococcus, in spu- 
tum, 11, 51 
Cytodiagnosis of ascitic fluid, 
700 
of cerebrospinal fluid, 684 
of pleural fluid, 700 



D 



Dahlia stain, 473 

Dare's haemoglobinometer, 443 

Day and night urine, 90 

Deen's test, 364 

Deficit of HC1, 348 

saturation, 348 
Degeneration, amyloid, urine 
in, 333 

anaemic, of red corpuscles, 
475 



Degenerations of red cells, 448 
Dehli boil, 673 
Delayed urea excretion, 311 
Dementia praecox, leucocytes 

in, 517 
Deneke's spirillum, 423 
Desmoid test, 354 
Desquamative nephritis, urine 

in, 32 7 
Desquamatory bronchial ca- 
tarrh, 69 
catarrhal pneumonia, 13 
of bowel, 397 
Determann's fluid, 526 
Deutero-albumosuria, 243 
Diabetes insipidus, 205 
mellitus, 198 

blood in, 649 
spinal fluid in, 688 
renal, 165, 551 
Diabetic coma, 202 

leucocytes in, 517 
Diacetic acid, 192 
Diagnosis, fibrin, 437 

functional renal, 309 
Diamines in urine, 257 
Diarrhoea, 387 

anaemia due to, 598 
of children, blood in, 645 
pancreatica, 392 
Diastase in stools, 397 

in urine, 160 
Diastatic activity of blood, 554 
Diazo-test, urine, 157 
Dicalcium phosphate sediment 

249 
Differential counting, 498 
Differentiated inner body of 

erythrocytes, 474 
Diffuse nephritis, acute, urine 

in, 329 
Digestion, gastric, 

products of, 356 
leucocytosis, 569 
Digestive power of pancreatic 

juice, 382 
Dilated stomach, anaemia due 
to, 598 
contents of, 367 
Diluting fluids, blood, 454 
Dilution test of urine, 312 
Dimethylamidoazobenzol, 345 
Dimethylamidobenzaldehyde 

test, 159 
Diptheria, 85 
bacillus, 37 
blood in, 513, 634 
Diplococcus lanceolatus in 
sputum, 32 
pneumoniae, 32, 33 
Dipylidium caninum, 417 
Diseases of kidneys, urine in, 

324 
Distomum buski, 414 
crassum, 414 
hepaticum, 414 
lanceolatum, 415 
rathouisi, 415 
Dittrich's plugs, 6 
Doune's pus test, 275 
Dried residue, of blood, 431 

determination of, 693 
Drigalski and Conradi's me- 
dium, 421 
Dropsical cells, 475 
Drugs, effect of, on red count, 

486 
Dry catarrh, 71 
Ducrey's bacillus, 300 
Dum dum fever, 673 
Duodenal ulcer, 373 
Dust in sputum, 12 
Dysentery, anaemia due to, 598 
bacilli of, 424 
stools in, 423 
Dyspepsia, anaemia due to, 582 
nervous, 369 



Easily split sulphates, 137 
Echinococcus disease of kid- 
ney, 306, 338 

of lung, sputum in, 
11, 51. 
Eclampsia, urine, 335 
Ectasis of stomach, 365 

hypertonic, 365 
Eel, vinegar, in urine, 307 
Egyptian chlorosis, 410 

haematuria, 307 
Ehrlich's classification of leu- 
cocytes, 494 
anemic degeneration, 475 
hemoglobinemic degener- 
ation, 449 
Elasticity of red cells, 447 
heat fixation, 464 
Methylene blue degener- 
ation, 47 7 
stains, 

dahlia, 473 
triacid, 471 
triple, 471 
Elastic tissue in gangrene, 77 
in sputum, 11, 57 
pseudo-, in sputum, 16 
Electrical conductivity, 311 
Emotional albuminuria, 226 
Emotional glycosuria, 166 
Emphysema, chronic, bron- 
chitis of, 71 
Empyema, 703 

leucocytosis in, 515 

perforating lung 80 

Encephalitis, acute, spinal fluid 

in, 689 
Enchondromata, 9 
Endocarditis, leucoyctes in, 

514, 632 
Endoglobular degeneration, 

448 
Endothelial cell leucocytosis 
519 
leucocytes, 450 
stains for, 469 
Enriching method of Schot- 

telius, 423 
Entamoeba buccolis, 51 
coli, 401 
dysenteriae, 401 
histolytica 51,|401 
Enteritis membranacea, 393 
Enterococcus, 360 
Enterogenous albumosuria, 

236 
Enteroliths in stools, 398 
Enthelmintha, 407 
Eosinophile cells, 497 
granules, 492 
of marrow, 505 
leucocytes, 451 
Eosinophilia, 508, 524 

after tuberculin reaction, 

523 
bronchitis, 11 
due to parasites, 523 
in asthma, 522 
in diseases of bone mar- 
row, 521 
in malignant disease, 524 
in skin disease, 522 
of genital organs, 523 
of haematopoietic organs, 

521 
of lymph glands, 522 
of spleen, 522 
of sympathetic nervous 

system, 524 
medicinal, 524 
parasitic diseases, 523 
physiological, 521 
post -febrile, 523 
Eosinophilic bronchitis, 11 
Epicritical polvuria, 91 
Epiguanin, 120 



716 



INDEX 



Epilepsy spina] fluid in, 688 
Episarken, 120 
Epiastaxis, Gull's renal, 237 
Epithelial casts, 264 

origin of, 269 

cells in prostatic fluid, 308 

in sputum, 11, 12, 68 

in stools, 397 

of urine sediments, 262 

filtration, 318 

tubes, 264 
Erosions, hemorrhagic, of gas- 
tric mucosa, 374 
Erysipelas, leucocytes in, 515 
Erythremia, 47 7 
Erythroblasts, 501 
Erythrocytes, see Red Blood 

Cells 
Erythrocytosis, 487 
Erythrodextrin, 186 
Esbach tubes, 216 
Essential albuminuria, 227 

renal hematuria, 237 
Ethereal sulphates, 136, 139 
Euglobin, 218, 219 
European hook worm, 410 
Eustrongylus gigas in kidney, 

306 
Ewald-Boas test breakfast, 

342 
Exercise, leucocytosis due to, 

519 
Exogenous uric acid, 115 
Extraneous structures in spu- 
tum, 1 1 
Exudates, 700 



Faeces, 382 (see Stools) 
False albuminuria, 225 
Famine fever, 676 
Farrant's mounting fluid, 273 
Fasciola hepatica, 415 
Fasciolopsis buski, 413 
Fasting stomach, contents of, 

341 
Fat, assimilation, 425 
in blood, 452, 561 
in bloody fluids, 694 
in stools, estimation of, 

392 
in urine, 209 
Fat globules 

in blood, 452 
in sputum, 13 
Fat-splitting ferment in gas- 
tric juice, 354 
in pancreatic fluid, 382 
Fatty acid crystals in sputum, 
17, 18 
acids in gastric juice, 

359 
acids in stools, estimation 

of, 390 
casts, 266 
cells in sputum, 13 
concretions in urine 279 
granules in leucocytes, 

494 
kidneys, urine of, 325 
stools, 390 
Febrile albuminuria, 231 
albumosuria, 236 
diseases, leucocytes in, 
512 
Fecal vomitus, 340 
Fehling's test, determination 
of sugar, 169 
test for glucose, 169 
Ferment, fat-splitting, in gas- 
tric juice, 356 
of urine, 159 
Fermentation in gastric juice, 

359 
Ferments in gastric juice, 354 
in stools, 397 
in urine, 159 



Ferrocyanide test, albumi- 
nuria, 213 
Fetal blood, 507 
Fetid bronchitis, 73 
Fever, albuminuria in, 325 

blood in, 487 
Fibres, muscle, in gastric juice, 
363 
in stools, 396 
Fibrin casts in sputum, 24 
in urine, 266 

coagula in sputum, 63, 67 
diagnosis, 431 
network in fresh blood, 

452 
structures in sputum, 9 
Fibrinogen, 

in body fluids, 693 
Fibrinous bronchitis, 73 

structures in sputum, 9 
Fibrinuria, 224 

Fibroid form of pulmonary tu- 
berculosis, 55 
Filaria bancrofti, 673 
demarquai, 676 
diurnia, 676 
embryos in urine, 306 
gigas, 676 
loa, 676 
megalhassi, 676 
nocturna, 673 
ozzardi, 676 
perstans, 676 
Filariasis, 673 
Finely granular casts, 265 

cells, 451 
Finkler and Prior's spirillum, 

423 
Fischer's test meal, 342 
Fish tuberculosis, 29 
Fistula, jejunal, 386 
Fixation of complement, 569 
of specific gravity of 
urine, 90, 312 
Fixing methods for smears, 

464 
Flagellata in cancer of stom- 
ach, 380 
in sputum, 51 
in stools, 403 
in urine, 306 
Flagellum stains, 282 
Flat worms, 413 
Fleischl haemoglobinometer, 

440 
Flexner Harris bacillus, 424 
Flu, sputum of 65, 

blood in, 515 
Fluid, body, analysis of, 693 
cerebrospinal, 678 
gastric, 339 
pancreatic, 340 
prostatic, 308 
Fluke, lung, 52 

Folin's acetone method, quan- 
titative determination, 
192 
ammonia method, 124 
creatinin method, 126 
urea method, 112 
uric acid method, 117 
Folin and Farmer's method for, 

nitrogen, 108 
Follicular tonsillitis, blood in, 

513 
Forceps for blood work, 447 
Foreign bodies in sputum, 11, 

78 
Form of stools, 387 
Formic acid in gastric juice, 

359 
Fractional study of gastric juice, 

351 
Fragments of cancer in gas- 
tric juice, 363 
of mucosa in gastric juice 

371 
of tissue in sputum, 17 



Fragments of tissue in sputum 
of abscess of lung, 78 
in sputum of gan- 
grene of lung, 77 
in urine, 274 
of tumors, in stools, 401 
in urine, 274 
in vomitus; 341 
Free HC1, determination of, 

345, 347 
Freezing point of urine, 310 
Fresh blood, study of, 445 
Fnedlander's bacillus, 34, 64 

formula, 157 
Fructose, 182 
Fuchsin, 281 
Fuchsinophilia, 476 
Fuchsinophilic cells, 501 
Functional albuminuria, 225 

renal diagnosis, 309 
Fungi in sputum, 9 
Furbinger's books, 297 
Furfurol test, 111 
Fusiform bacillus, 41 



Gabbett's methylene blue, 26 
Gall-sand, 398 

Gall-stone colic, blood in, 656 
Gall-stones, 398 
pseudo-, 398 
Gamete, 662 
Gangrene of lung, leucocytes 

in, 517 
sputum of, 77 
Gas bacillus, 296 
Gastric contents, 339 
achylia, 372 
acidity, 344 
ammonia in, 359 
bacteria in, 362 
bases of, 359 
blood in, 363 
cancer, 364 
cancer ferments in, 

357, 377 
cells of, 361 
digestion products, 

356 
epithelial cells in, 

361 
fat splitting ferment, 

356 
fermentation, 359 
fractional determin- 
ation of, 351 
fragments of mucosa, 

363 
hydrochloric acid, 

344 
in acute gastritis, 369 
in atrophy of mucosa, 

372 
in cancer, 375 
in chronic gastritis, 

370 
in hyperacidity, 367 
in hypersecretion, 367 
in nervous dyspepsia, 

369 
in pernicious anemia, 

350 
in ulcer, 373 
infusoria in, 362 
lactic acid, 357 
lipase, 356 
moulds, yeasts, sar- 

cinae in, 363 
mucus in, 371 
muscle fibres in, 361 
occult blood, 364 
organic acids, 359 
pus in, 362 
rennin, 356 
sarcinse in, 360 
starch digestion, 357 



INDEX 



717 



Gastric tumor, fragments inV 
365 
yeasts in, 360 

diagnosis without tube, 
366 

digestion, products of , 356 

microscopic examination 
of, 363 

motility, 341, 364 

mucosa, atrophy of, 372 

pepsin, 354 

sediment, 360 

ulcer, 373 
Gastritis acida, 371 

acuta, 369 

chronica, 370 

interstitial purulent, 370 
Gastritis, acute and chronic 
anaemia due to, 598 j 

phlegmonosa, 362, 370 

purulentia, 370 
Gastro-intestinal disorders, an- 
aemia due to, 599 
Gastrosuccorrhcea, 369 
Gastroxynsis, 368 
Gelatine, 286 

General paresis, blood in, 517, 
647 
spinal fluid in, 685 
Genitalia, eosinophilia in dis- 
eases of, 523 

infection of, 294 
Gentian violet, 33, 281 
Geraghty and Rowntree's test, 

319 
Gerhardt's test, diacetic acid, 

193 
German measles, blood in, 633 
Giant cells of bone marrow, 

507 
Giant phagocytes, 507 
Giemsa's stain, 301 
Gigantocytes, 475, 606 
Glanders of lung, 66 
Glassy casts, 267 
Globular decolorization, 448 

richness, blood, 608 
Globulin in body fluids, 693 

serum, in urine, 218 

in spinal fluid, 681 
Glossina morsitans, 673 
Glucose, 162 

quantitative test for, 175 

to remove, 196 
Gluzinski's test, 380 
Glycerine agar, 285 

bouillon, 29 
Glycocholic acid, 155 
Glycogen in urine, 186 
Glycoproteids, 694 
Glycosuria, 162, 164 

eamylo, 162 

emotional, 166 

esaccharo, 162 

renal, 165 
Glycuresis, 164 
Glycuronic acid, 162, 206 
Gmelin's test, 151 
Gonococcus, 294 
Goetch's test, 551 
Gonorrhea, complement fixa- 
tion for, 584 
Gonorrhceal, 293 

arthritis, leucocytosis in, 
516 

cystitis, 293 

threads, 273 
Gout, leucocytes in, 516 
Gouty albuminuria, 230 
Gower's hsemoglobinometer, 

441 
Gram's stain, 38 
Gram-negative organisms, 38 
Gram-oositive organisms, 38 
Granular casts, 265 

cells of bone marrow, 503 

cells in prostatic fluid, 308 I 



Granular masses of Schultze, 

452 
Granules, acidophile, 492 

amphophilic, 492 

basophile, 492 

of Grawitz, 479 

eosinophil, 492 

fatty, '494 

Grawitz, 479 

haemokonien, 451 

in red cells in malaria, 
477 

Mastzell, 492 

Morris, 450 

neutrophile, 493 

of leucocytes, 491 

of lvmphocytes, 493 

of red cells, 477, 479 

oxyphilic, 492 

perinuclear, 493 

pigment, in sputum, 4, 12 

sago, in sputum, 2 

Vaughan's, 449 
Grass bacillus, 29 
Gravel-forming enteritis, 400 
Grawitz granules, 479 
Grawitz's unripe cell, 505 
Green sputum, 4, 62 

vomitus, 340 
Grinder's rot, sputum of, 5 
Grippe, 65 
Groups blood, 588 
Growth, anaemia of, 645 
Gruber-Widal test, 564 
Guaiac test, 241, 394 
Guanin, 120 

Gull's renal epistaxis, 237 
Gum, animal, in urine, 187 
Gunning's test, acetone, 189 

method for nitrogen, 107 
Gunzburg's reagent, 345 



hi 



Hammarsten's bile test, 153 
Hammerschlag's method of 
determining specific gravity 
of blood, 429 
Hanot's disease, blood in, 657 
Hard chancre, organism of, 

300 
Haser's coefficient, 94 
Hasting's stain, 467 
Hayem's fluid, 454, 527 
Hay's bile acid test, 156 
Healthy, albuminuria of, appa- 
rently, 22 7 
Heart disease, blood in, 65 7 

effect of, on count of 
reds, 657 
Heat test, albumin, 210 
Heating smears, 464 
Hehner-Maly method, organic 

acids, 359 
Heller's test, albumin, 212 
blood, 240 
quantitative, 221 
Hemagglutinins, 588 
Hemameba leukaemiae magna, 
62 7 
parva, 627 
malariae, 662 
virax, 660 
Hematoblast, 524 
Hematochyluria, 209, 676 
Hematocrit, 460 
Hematogenous albuminuria, 
231 
albumosuria, 235 
jaundice, 149 
Hematoidin in sputum, 18 
in urine sediment, 253 
Hematopoietic organs, 591 

eosinophilia in dis- 
eases of, 521 
Hematoporphyrin, 95, 242 
Hematoporphyrinemia, 243 
Hematoporphyrinuria, 242 



Hematozoon falciparium, 664 
Hematuria, 97, 237 

angioneurotic, 23 7 

Egyptian, 307 

essential, 23 7 

renal, 23 7 

terminal, 296 
Hemin test, 240 
Hemin test, blood in urine, 

240 
Hemoblast, 500 
Hemocytometers, 452 
Hemoglobin, 439 

casts, 265, 267 

estimation of, 439 

in sputum, 4, 13 

in urine, 340 

in urine sediment, 253 
Hemoglobinaemia, 528 
Hemoglobinaemic degenera- 
tions, 449 
Hemoglobinometers, 439 
Hemoglobinuria, 238 

paraxysmal, 239, 528 
Hemoglobmuric nephritis, 328 
Hemokonien granules, 451 
Hemolytic anemia, 602 
Hemolytic system, 574 
Hemometer, 441 
Hemophilia, 433; renal, 237 
Hemoptysical phthisis, 58 
Hemoptysis, 58, 81 

calculosa, 10 

non-tuberculous, 5 

parasitic, 52 
Hemorrhage, anaemia due to, 
595 

gastric, 364 

in bronchiectasis, 76 

leucocytosis due to, 518 

occult, in stools, 364 

pulmonary, 81 
Hemorrhagic erosions of gas- 
tric mucosa, 374 

infarct of lung, 83 

nephritis, 246 

pleurisy, 704 

pneumonia, 62 

sputum, 3 
Hemorrhagic erosions of 

stomach, 374 

infarct of lung, 83 
Hemosiderin in urine, 261, 

granules, 451 
Hepatogenous albumosuria, 
236 

jaundice, 148 
Hereditary albuminuria, 230 
Herzfehlerzellen, 4, 13 
Hess's viscosimeter, 437 
Hetero-albumose, 253 
Hetero-albumosuria, 233 
Heterotricha, 416 
Heteroxanthin, 120 
Hippuric acid, 253 

test, 323 
Hodgkin's disease, 628 
Homogentisinic acid, 98, 208 
Hook worms, 410 
Horismoscope, 212 
Howell's method of determin- 
ing coagulation time, 436 
Hunger diabetes, 164 
Huppert's bile test, 153 
Hyaline, 661 

casts, 267 
Hydatid disease of kidney, 338 
Hydremia, 591 
Hydrobilirubin, 150 
Hydrocele fluid, 708 
Hydrocephalus spinal fluid, 

688 
Hydrochinone, 98, 148 
Hydrochloric acid in gastric 
juice, 344 
absolute amount, 349 
bound, 344 
deficit, 348 



718 



INDEX 



Hydrochloric acid, free, 344 

determination of, 347 
quantitative deter- 
mination of, 345 

Hydrochloric acid, tests for, 
345 

total, 348 

Hydrogen disulphide in gas- 
tric contents, 360 

ion concentration of blood 
532 

Hydrogen sulphide in urine, 
140 

Hydronephrosis, 338 

Hydroxy butyric acid, 194 

Hygrometry, 431 

Hymenolepsis nana, 417 

Hyperaciditas hydrochlorica, 
367 

Hyperacidity, 367 

Hyperalbuminemia rubra, 608 

Hyperchlorhydria, 367 

Hyperglycemia, 549 

Hyperlipemia, 260 

Hypermeability renal, 321 

Hypermotility, 364 

Hypersecretion, 367 

Hypertonic ectasis, 365 

Hypocythasmia, 590 

Hypoglycemia, 549 

Hypoglycosuria, 324 

Hypomotility, 365 

Hypoplastic anaemia, 591 

Hypostatic albuminuria, 320 
pneumonia, 65 

Hyposthenuria, 312 

Hypoxanthin, 120 



Ideopathic fibrinous bronchitis 
73 
pentosuria, 184 
Immature nucleated reds, 500 
Inanition, anaemia of, 597 

blood in, 658 
Index, color, 490, 591 

volume, 491 
Indifferent lymphoid cells, 506 
Indigo calculi, 144, 279 
carmine test, 318 
sediment, 253 
Indigo-blue in urine, 145 

' red in urine, 147 
Indoxyl sulphate in urine, 98, 

143 
Indurative nephritis, urine in, 
330 
pneumonia, sputum of, 65 
Infarction of kidney, urine in, 
336 
of lung, sputum in, 
83 
Infectious disease, cause of 

anasmia, 599 
Infectious, latent anemia due 

due to, 598 
Infectious nephritis, urine in, 

291 
Inflammatory leucocytosis, 

512 
Influenza, bacillus of, 34 

leucocytes in, 515 
sputum of, 65 
Influenzal meningitis, 690 
Infusoria in cancer of stom- 
ach, 380 
in gastric contents, 362, 

363 
in intestine, 401 
in sputum, 50 
in stools, 406 
Inner body of Lowit, 474 
Inorganic acids in urine, 129 

bases in urine, 129 
Inoscopy, 702 
Inosite, 186 

in body fluids, 698 



Inosite in diabetes insipidus, 
206 

in urine, 186 
Insanity, blood in, 647 
Insipidus, diabetes, 205 
Insufficiency, glomerular and 
tubular, 312 
motor, of stomach, 365 
Insular nephritis, 230 
Intermediate forms of nucle- 
ated reds, 501, 607 
Intermittent albuminuria, 22 7, 

330 
Interstitial gastritis, purulent, 
370 
nephritis, urine in, 329 
pneumonia, sputum of, 65 
Intestinal cancer, blood in, 650 
concretions, 398 
contents, 382 
Intestinal obstruction, leuco- 
cytes in, 514 
parasites, 401 

blood in, 642 
cause of anaemia, 600 
sand, 400 
test meals, 385 
worms, 401 
Intestine, contents of, 385 

fat splitting ferment, 382 
lipase, 382 
motility of, 385 
motility, 385 
pancreatic fluid, 340, 382 
trypsin, 382 

tuberculosis of, blood in, 
638 
Intraligamentous ovarian 

cysts, fluid of, 708 
Iodine reaction in blood, 524 
Iodophilia, 524 
Iron in urine, 142 
Irritation forms of luecocytes, 

498 
Isomaltose, 162, 187 



Jaffe's test for creatinin, 126 
for indoxyl sulphate, 
145 
Jaundice, catarrhal, blood in, 
656 
gastric contents in, 351 
haematogenous, 149 
hepatogenous, 148 
sputum in, 4 
toxaemic, 149 
toxic, blood in, 656 
Jejunal fistula, 386 
Joints, tuberculosis of, blood 

in, 638 
Juice, gastric, acidity of, 344 
Justus's test, 654 



K 



Kahler's disease, 233 
Kala azar, 673 
Kaufmann's bacillus, 379 
Kemp's fluid, 526 
Kidney, abscess of, urine in, 
336 
acute nephritis, blood in, 

654; urine in, 327 
amyloid degeneration of, 

urine in, 333 
arteriosclerosis of, urine 

in, 332 
atrophy of, urine in, 335 
cancer of, 336 
chronic passive congestion 

of, urine in, 327 
cloudy swelling of, urine 

in, 326 
congenital cystic, blood 
in, 656; urine in, 335 



Kidney, diseases of, urine in 
324; 

blood in, 654 
echinococcus disease, 338 
fatty, urine in, 326 
infarction of, 336 
large white, urine in, 329 
nephritis, blood in, 654 

urine in, 32 7 
parasitic diseases of, 338 
senile atrophy of, urine 

in, 325 
stone in, 2 76 
suppurative nephritis, 336 
tuberculosis of, 336 
blood in, 638 
Kjeldahl method, 107 
Klebs-Loffler bacillus, 37 
Kulz sign, 272 



Labor, albuminur.a of women 

in, 227 
Lactic acid, 694 

bacillus, 288 

in body fluids, 695 
in cancer of stomach 

377 
in gastric juice, 357 
quantitative deter- 
mination of, 359 
Lactorrhea, 659 
Lactose, 182 

test of Schlayer, 323 
Laiose, 182, 187 
Lamblia intestinalis in stools, 

404 
Landsteiner's blood groups, 598 
Landwehr's animal gum, 189 
Large lymphocytes, 505 

mononuclears, 450, 495 

white kidney, urine in, 
331 
Latent constipation, 387 
Layer formation, sputum, 5 
Lead in urine, 143 

poison, cause of anaemia, 
601 

cause of stippled cells, 481 
Lecithin, 556 

in body fluids, 694 

globules in prostatic fluid, 
308 
Legal's test, 188 
Leishman- Donovan bodies, 

672 
Leishmaniasis, 672 
Lennec's perles, 66 
Leprosy, bacillus of, 29, 31 

blood in, 657 
Leptodera intestinalis, 413 

stercoralis, 413 
Leptothrix group, 32 

innominata in sputum, 32 

maximus buccalis, 32 
Leucin, 696 

in body fluids, 695 

as sediment, 254, 256 

in sputum, 18 
Leucoblast, 505 
Leucocytes, 450, 491, .494 

coarsely granular, 45 1 

counting, 461 

endothelial, 450 

eosinophile, 451 

finely granular, 451 

granules of, 491 

large mononuclears, 450 

lymphocytes, 450 

mastzellen, 450 

pigmented, 451 

polymorphonuclears, 451 

small mononuclear, 450 

transitionals, 450 . 
Leucocytosis, 508; abscess, 516 

active, 520 

acute bronchitis. 516 



INDEX 



719 



Leucocytosis, acute 'cerebro- 
spinal meningitis, 513 
acute fibrinous pleurisy, 

515 
acute follicular tonsilitis, 

513 
acute poliomyelitis, 514 
acute ulcerative endocar- 
ditis, 514 
agonal, 518 
anthrax, 516 
appendicitis, 644 
arthritis, 516 

articular rheumatism, 513 
bronchiectasis, 516 
bronchitis* 516 
cerebrospinal meningitis, 

513 
.children, 520 
cholera, 514 
chronic bronchitis, 516 
dementia praecox, 517 
diabetic coma, 517 
digestion, 519 
■diphtheria, 513 
empyema, 515 
endocarditis, 514 
endothelial cell, 519 
-erysipelas, 514 
exercise, 520 
febrile diseases, 512 
fetid bronchitis, 516 
follicular tonsillitis, 553 
gangrene of lungs, 516 

gout ,516 
liydronephrosis, 516 _ 

inflammations and febrile 

diseases, 512 
influenza, 515 

intestinal obstruction, 514 

lobar pneumonia, 513, 640 

malignant disease, 517 

Mastzell, 528 

medicinal, 519 

meningitis. 514 

mixed, 520 

myxcedema, 514 
•of eosinophiles, 521 

of large mononuclears, 

519 
•of new-born, 511 

paresis, 517 

passive, 520 

pelvic abscess, 5 1 7 

perirenal abscess, 516 

pleurisy fibrinous, 515 
with effusion, 515 

physiological, 509 

pneumonia, 513, 640 

poliomyelitis, 514 

post-hemorrbagic, 518 

post operative, 517 

pregnancy, 510 

pyelitis, 516 

pyelonephritis, 516 

pyelonephrosis, 516 

pyogenic inflammations, 
515, 517 

rabies, 514 

renal calculus, 516 

rheumatism, 513 

scarlet fever, 633 

smallpox, 634 

tonsillitis, 513 

uraemia, 517 

whooping-cough, 513 

women, 511 
Leucopenia, 521 
Leukaemia, 616 

acute, 62 7 

lymphatic, 616, 623 

mixed, 615, 628 

"parasite, " 626 

spleno-myelogenous, 616 
Leukanaemia, 629 
ILevulose, 181 
Levulosuria, 181 



Leyden (Charcot-) crystals in 
blood, 620 
in sputum, 18 
in stools, 398 
Lientery, 396 

Lilienfeld's nucleohiston, 223 
Lipemia, 556 

Lipase, in pancreatic juice, 382 
in stomach, 354 
in urine, 160 
Lipoids of blood, 555 
Lipuria, 209, 259 
Litmus milk, 286 
Liver abscess, blood in, 656 
acute yellow atrophy, 

blood in, 65 7 
cirrhosis, blood in, 657 
diseases, blood in, 656 
perforating through lung, 
79 
Lobar pneumonia, (see pneu- 
monia) 
Locomotor ataxia, spinal fluid, 

in, 686 
Loffler's blood serum, 37 

methylene blue, 38 
Lowit's inner body, 474 

organism, 627 
Lowy, alkalinity of blood, 537 
Lues, blood in, 653 

congenital, blood in, 646 
of lungs, sputum in, 84 
organism of, 300 
serum diagnosis of, 575 
urine in, 329 
Luetic nephritis, 329, 333 
Lung, abscess of, blood in, 516 

sputum of, 78 
Lung, actinomycosis of, spu- 
tum of, 45 
blastomycosis of, 42 
cancer of, sputum of, 83 
chronic passive conges- 
tion of, sputum of, 83 
echinococcus of, 11, 51 
fluke, 52 

gangrene of, blood in, 
517; sputum of, 77 
glanders of, 66 
liver perforation through, 

79 
lues of, 84 
moulds of, 45 
oedema of, 80 
stones, 9 

tuberculosis of, blood in, 
634; sputum in, 54 
Lutein, 152 
Lymphadenoid leukemia, 616, 

625 
Lymph glands, eosinophilia in 
diseases of, 524 
tuberculosis of, 636 
Lymphatic leukaemia, 623 
Lymphemia, 616, 623 
Lymphocytes, 494 

large of marrows, 505 
granulations of, 493 
Lymphocytosis, 519 
of infants, 520 
Lymphomatosis, alymphemic, 

522 
Lymphuria, 209 

M 

Macrocytes, 475 

Macrogamete, 662 

Macrophages, 452 

Macroscopic constituents, spu- 
tum, 6 

Macroscopy of stools, 415 

Magnesium, 140; phosphate in 
urine, 249 

Malaria, albuminuria in, 326 
blood in, 629, 659 
of children, 646 



Malachite green bouillon, 422 

stain, 26 
Malarial parasites, asstivo-au- 
tumnal, 664 
quartan, 662 
tertian, 660 
Mai de Caderas, 671 
Malignant diseases, blood in, 
517, 648 
eosinophilia in, 524 
gastric juice in, 351 
leucocytes in, 517 
of lung, sputum of, 
83 
Maltose, 187 
Manure bacillus, 29 
Maragliano's endoglobular de- 
generation, 448 
Marguerite form, 663 
Marrow, bone, 500 
Marshall's urease method, 

112 
Massea's spirillum, 423 
Masturbators, albuminuria of, 

227 
Mastzell granules, 492 
leucocytosis, 519 
Mastzellen, 450, 497, 505 
Mature nucleated reds, 500 
Mayer's solution, 467 
McCrudden's method, faeces 

examination, 386 
McGowan's method stool ex- 
amination, 434 
Mcjunkin's stain, 469, 496 
Meals, test, 342 

Dock's 342 . 
Ewald-Boas, 342 
Fischer's, 342 
for intestinal condi- 
tions, 385 
lactic acid, free, 633 
Mosenthal's, 313 
O'Hare's, 314 
renal, 342 
Riegel's, 342 
Measles, blood in, 633 

German, blood in, 633 
Media for bacteria, 35, 285 
Mediastinal growths, 84 
Medicinal eosinophilia, 524 

leucocytosis, 518 
Medicines, color of urine due 

to, 99 
Megaloblasts, 501, 606 
Megalocytes, 475 
Megalogastria, 365 
Megalokaryocyte, 506 
Megastoma entericum, 362 
Melanin, 97, 155 

as urine sediment, 253 
Melanogen, 155 
Melituria, 187 
Mellitus diabetes, 198 
Melting point of crystals, de- 
termination of, 173 
Membranous enteritis, 393 

ureteritis, 221 
Meningitis, cerebro-spinal leu- 
cocytes in, 513 
spinal fluid in, 690 
tuberculous, spinal fluid 
in 690, blood in, 639 
Meningococcus, 701. 
Mental disease spinal fluid in 

588 
Merozoite, 659 
Metamyelocyte, 504 
Metalbumin, 706 
Methemoglobin, 242 
Methemoglobinemia, 528 
Methylalcohol, 464 
Methylene-blue, degeneration 
of Ehrlich, 477 
Eosin stain, 467 
Gabbett's, 26 
Loeffler's, 38, 280 
saturated aqueous, 281 



720 



INDEX 



Methylene-blue stains, 2, 467 
test in urine, 318 
urine, 99 
Methyl-violet test, 345 
Methylxanthin, 120 
Metrocytes, 504, 507, 605 
Mett's method for pepsin de- 
termination, 354 
Microblasts, 502, 607 
Microchemistry of urine, 244 
Micrococcus aureus, 19 

catarrhalis, in sputum, 22 
intracellularis meningiti- 
dis, 690 
pneumoniae, 32, 64 
cultivation, 33 
types, 33 
tetragenus in sputum, 41 
Microcytes, 474 
Microgamete, 662 
Mjcrogametocyte, 662 
Microscopic constituents of 
sputum, 11 
examination of gastric 

contents, 363 
of stools, 397 
Miescher's hasmoglobinometer, 

.439 
Miliary tuberculosis, blood in, 
637 
sputum of, 55 
Milk and butter bacillus, 29 
Milk.curds in stools, 396 

litmus, 286 
Milk examination of, 699 
Milk-white zone, 668 
Millian's method, 434 
Mineral acidity of urine, 111 
Miner's anemia, 410 
Minimal albuminuria, 230 
Mixed leukemia, 628 
leucocytosis, 519 
Mceller's bacilli, 29 
Monocercomonas hominis, 404 
Monont, 659 

Mononuclears, eosinophile, 
498 
large, 495 

increase of, 519 
neutrophile, 498 
small, 494 
Morner's body, 219, 222 
Morning sputum, 2 

star crystals in urine, 246 
Mosquito-cycle of malarial 

parasite, 665 
Moss blood groups, 588 
Motility of intestine, 385 
of stomach, 341, 364 

determination of, 381 
ectasis, 365 
hypermotility, 365 
motor insufficiency, 

365 
salicylic-acid test, 

366 
Motor insufficiency of stom- 
ach, 365 
Moulds in gastric contents, 
363 
in sputum, 45 
in stools, 418 
in urine, 306 
Much's granules, 26 
globules, 298 
m sputum, 54 
Mucinuria, 221 
Mucoid in body fluids, 695 

sputum, 3 
Mucopurulent sputum, 3 
Mucor corymbifer, 46 
mucedo, 46 
pusillus, 47 
racemosus, 47 
rhizopodiformis,47 
spetatus, 47 



Mucosa, atrophy of, 3 72 

fragments of, in gastric 
contents, 371 
Mucous colitis, 393 

corpuscles, 68, 261 

sediment, 261 

threads in urine, 274 
Mucus in stomach, 371 

in stools, 392 

in urine, 220 

in vomitus, 340 

sediment, 261 
Miiller's blood dust, 491 
Mumps, leucocytes in, 514 
Murexid test, 116, 247 
Muscle fibres in gastric con- 
tents, 361 
in stools, 396 
Myelemia, 509, 616 
Myelin in sputum, 2, 13 
Myeloblast, 504 

of Nageli, 505 
Myeloc3^tes, eosinophile, 499 

Cornil's, 619 

neutrophile, 497, 504 
Myelogenous leukemia, 615 
Myelopathic albumosuria, 233 
Myxcedema, blood in, 658 

leucocytes in, 514 



N 



Naeglaei's myeloblast, 505 
Nakayama's test for bile, 153 
Necator americanus, 410 
Necrotic tissue fragments in 

sputum, 6 
Neisser's stain, 38 
Nematode worms in urine, 307 
Nephritis, acute, 327 

albuminuria of, 232 
blood in, 655 
chronic diffuse, 329 
indurative, 330 
interstitial, 332 
parenchymatous, 329 
desquamative, 327 
diffuse, 32 7 

parenchymatous, 329 
haemoglobinurica, 328 
hemorrhagic, 320 
indurative, 330 
infectious, 291 
interstitial, 329 
insular, 130 
■ luetic, 233, 329 
non-indurative, 329 
of cholera, 329 
parenchymatous, 326 
purulent, 336 
subacute, 329 
suppurative, 336 
unilateral, 335 
urine cultures in, 290 
Nervous disease, blood in, 641 
dyspepsia, 369 
form of albuminuria, 23 7 
system, sympathetic, dis- 
eases of, eosinophilia in, 
524 
Nessler's solution, 109 
Neutral magnesium phosphate 

crystals, 249 
Neusser's granules, 493 
Neutral sulphates, 137 
Neutrophile granules, cells, 
492, 496 
myelocytes, 504 
Neutrophiles, small, 498 
New-born, albuminuria of, 227 

leucocytosis of, 511 
New growth in kidney, urine 
in, 336 
in mediastinum, spu- 
tum in, 84 
Night and day urine, 90 



Nitric acid in urine, 140 

test for albumin, 211 
for urea, 111 
Nitrogen of blood, 539 
of sputum, 19 
of urine, 104 

determination of, 107 
Nitrogenous bodies of urine,. 
104 
blood plasma, 540 
Nitrous acid, 140 
Noguchi test, 577 
Non-indurative nephritis, 329 
Non-proteid nitrogen of blood, 

540 
Normal persons, sputum of, 2 
Normoblasts, 501 

in pernicious anaemia, 606 
Nubecula, 100, 261 
Nucleated reds, 448, 500, 605 
ancestral, 504 
Howell's mature and 

immature, 500 
intermediates, 501 
Nuclei of reds, changes in, 502 

fate of, 502 
Nuclein bases, 120 
Nucleinic acid, 162, 222 
Nucleo-albumin, 211, 213, 219, 

221 
Nucleoabluminuric, 223 
Nucleohiston, 223 
Nucleoid, 474 
Nummular sputum, 58 
Nutrient agar, 285 
Nutrition, effect of, on red 

count, 484 
Nycturia, 90 
Nylander's test, glucose, 170 



Obermayer's test for indoxyl, 

145 
Obstruction of intestine, blood 

in, 514 
Occult hemorrhage in, gastric 
contents, 364 

stools, 394 
Ochronosis, 98 
Odor of sputum, 5 

of urine, 99 
(Edema, chloride retention in, 

130 
GMema of lungs, sputum of, 80 
Oesophagus, cancer of, blood 

in, 651 
Oidium albicans in sputum, 44 
in stools, 418 

coccidioides, 44 
Oligemia, 571 

serosa, 591 

sicca, 591 
Oligochromemia, 445, 590 
Oligocythemia, 482, 590 
Oligoplasma, 591 
Oliguria, 90 
Oocyst, 665 
Olkinet, 665 

Operation, leucocytosis after, 
517 

red count after, 487 
Oppler-Boas bacillus, 362, 378 
Opsonic index, 568 
Opsonins, 567 
Orcin test, 185 

Organic acids in gastric con- 
tents, 104 
Organic acidity of urine, 104 
Organized sediments, 244 
Oriental sore, 673 
Orthochromatic cells, 501 
Orthostatic albuminuria, 227 
Orthotic albuminuria, 227 
Osier's disease, 488 

protoleucocyte, 505 
Osteoclasts, 507 



INDEX 



721 



Osteoma of lung, 9 
Ovarian cysts, fluid of, 706 
Over work, anemia due to, 597 
Oxalate, calcium, crystals in 

sputum, 18 

in urine, 250 

stones, 278 
Oxalic acid, 118, 252 
Oxaluria, 250 
Oxybutyric acid, 194 
Oxyphilic granules, cells, 492 

cells, 501 
Oxyproteinic acid, 128 
Oxyuris vermicularis, 408 



Palpation, albuminuria due 

to, 231 
Pancreatic diarrhea, 392 

disease, stools in, 392, 
424 
fluid, 340, 382 
secretion in vomitus, 351 
stones, 398 
Paracellaire albuminurie, 230 
Paracresol, 148 

Paragonimus westermani, 52 
Paramecium coli, 406 
Paramucin, 706 
Paranucleoproteid, 694 
Parasites, anaemia due to, 600 
animal, in lung, 50 

in urine, 306 
eggs, preservation of, 426 
eosinophilia in, 523 
intestinal, 401 
plant, in sputum, 18 
in stools, 418 
Parasitic chyluria, 260 

diseases of kidney, 338 
haemoptysis, 52 
Paratyphoid bacillus, 287 
Paraxanthin, 120 
Parenchymatous nephritis 

acute, urine in, 327 
chronic, urine, in, 329 
Paresis, blood in, 518, 648 

spinal fluid in, 685 
Parovarian cyst fluid, 708 
Paroxysmal hemoglobinuria, 
528 
hematoporphysinuria, 242 
polyuria, 91 
Pavement-epithelial cells in 

sputum, 11 
Pavy's disease, 229 
Pea soup stools, 420 
Penicillium glaucum, 50 

nummula, 50 
Pentoses, 162, 173, 183 
Pentosuria, 183 
Pepsin, 354 

quantitative method, 354 
in urine, 159 
Peptonuria, 235 
Perforating empyema, lung, 
80 
pleurisy, serous, 80 
Pericardial fluid, 704 
Perinuclear granules of Neus- 

ser, 493 
Periodic polyuria, 91 
Perirenal abscess, leucocytes 

in, 516 
Peritoneal fluid, analysis of, 

700 
Peritonitis, tuberculous, blood 

in, 638 
Perles of Lasnnec, 66 
Permanganate index, 683 
Permeability, renal, 318 
Pernicious anaemia, 602 

gastric juice in, 350 
Pertussis, leucocytes in, 513 
organism of, 37 
sputum of, 66 
Pessary forms, 447 



Pettinkofer's reaction, 156 
Petroff's medium, 27 
Pfeiffer's bacillus, 34 
Pfluger's tubules, cysts of, 707 
Phagocytic cells of blood, 496 
Phagocytosis, 568 
Pharyngomycosis, 42 
Phenol in urine, 144 
Phenolphthalein, 346 
Phenolsulphonepthalein test, 

319 
Phenolsulphuric acid, 148 
Phenylhydrazin test for glu- 
cose, 172 
Phlegmonous gastritis, 370 
Phlorizin test, 323 
Phloroglucin reaction, 184 
Phosphate concretions, 278 

crystals in sputum, 18 

sediment, 248 
Phosphates, calcium, 249 

in urine, 248, 133 

magnesium, 249 

triple, 248 
Phosphatic diabetes, 134 
Phosphaturia, 100, 101 
Phosphaturique albuminurie, 

230 
Phosphorus-containing pro- 

teids, 694 
Phosphorus poisoning, red 

count, 487 
Phthisis, blood in, 634 

haemoptisical, 458 

melanotica, 13 

renal, 336 

stone-cutters, 5 
Physiochemical renal test, 310 
Physiological albuminuria, 224 
227 

eosinophilia, 522 

leucocytosis, 509 

variation in red count, 
483 
Picini's fluid, 526 
Pigment, bile, in sputum, 4 
in urine, 97 

blood, in sputum, 4, 13 

coal, in sputum, 4 

of urine, 143 
Pigmented cells in blood, 630 

in sputum, 13 

of marrow, 506 
Pigments of urine, 94 
Pin worms, 408 
Piria's test, 255 
Piroplasmosis, 672 
Placental cells, 452 
Plague pneumonia, 65 
Plant parasites in sputum, 18 
in stools, 418 
in urine, 279 
Plasmodium praecox, 664 

vivax, 660 
Plastic bronchitis, sputum of, 

73 
Platelets of blood, 452 
Plaut's angina, 87 
Plehn's granules, 477 
Plethora vera, 591 
Pleural fluid, analysis of, 700 
cytodiagnosis of, 700 
Pleurisy, acute fibrinous, leu- 
cocytes in, 515 

serous, perforating, spu- 
tum in, 80 
Pleurisy, with effusion, leuco- 
cytes in, 515 
Plugs, Dittrich's, 6 

prostatic, 309 
Pneumaturia, 305 
Pneumococcus, 32, 64 

mucosus, 22 

meningitis, 690 
Pneumokoniosis, 85 
Pneumoliths, 10 
Pneumonia, abscess of lung, 78 
acute lobar, sputum of, 60 



Pneumonia, albuminuria in, 326 

aspiration, 65 

blood in, 513, 640 

broncho-, blood in, 642 
sputum of, 65 

chlorides in, 129 

chronic, sputum in, 60 

congestion, 62 

croupous, sputum in, 60 

desquamatory catarrhal, 
13 

green sputum of, 62 

hypostatic, 65 

interstitial, 65 

plague, 65 

serous, 62 

sputum of, 60 

subacute, sputum in, 65 

tuberculous, 55 
Pneumonomycosis aspergillina, 

49 
Pneumotyphoid fever sputum, 

of, 65 
Poikilocytes, 446, 605 
Poisons, anaemia due to, 600, 

598, 602 
Polariscope, 174 
Poliomyelitis leucocytosis in, 
514 

spinal fluid in, 692 
Polychromatophilia, 475, 501 
Polychrome methylene-blue 

stains, 467, 470 
Polycythemia, 482, 487 
Polymastigina, 403 
Polymorphokaryocyte, 506 
Polymorphonuclear leucocytes, 

451, 497 
Polyplasma, 591 
Polyps in stools, 401 
Polyuria, 90; epicritical, 91 

paroxysmal, 91 

periodic, 91 
Polyvalent antigens, 584 
Poor, anaemia of the, 597 

food, anaemia due to, 597 
Post-febrile eosinophilia, 523 
Post-hemorrhagic anaemia, 594 

leucocytosis, 518 
Post-operative leucocytosis, 

517 
Postural albuminuria, 227 
Potassium ferrocyanide test 
for albumin, 213 

in urine, 142 

iodide test, 148 
Pregnancy, ammonia in, 123 

leucocytosis of, 510 

red cells in, 484 
Pregnant women, albuminuria 

of, 227 
Pregoutteuse albuminurie, 230 
Preservation of sediments, 243 

of urine, 88 
Primary anasmia, 602 

pernicious anaemia, 602 
Progressive pernicious anae- 
mia, 602 
Proliferating cysts, Pfluger's 

tubules, 707 
Promyelocyte, 504 
Prostatic fluid, 308 

plugs, 309 
Prostatis, 297 
Prostatorrhcea, 298 
Proteids in body fluids, 694 

of urine, 210, 217 
Proteins to remove from blood, 

196 
Proteus bacillus, 288 

cystitis, 293 

mirabilis, 288 

vulgaris, 288 

zenkeri, 288 

zoffii, 288 
Protoleucocytes, 505 
Protomonadina, 405 



46 



722 



INDEX 



Protoryxomyces coprinarius, 

404 
Protozoa, 401 

stains for, 470 
Prune-juice sputum, 63 
Pseudo-casts, 269 
Pseudochlorosis carcinomatosa, 
649 

tuberculosa, 635 
Pseudochylous fluids, 70S 
Pseudodiptheria bacilli, 86 
Pseudo-gall-stones, 398 
Pseudoglobulin, 218 
Pseudoleucocytosis, 517 
Pseudoleukaemia, 628 

of children, 646 
Pseudolymphocytes, 498 
Pseudomucin, 706 
Pseudonucleation, 448 
Pseudoparasites in red cells, 

448 
Pseudophthisis calculosa, 10 
Pseudotuberculosis organisms 
of, 30, 44 

moulds in, 49 
Pseudovacuolization of red 

cells, 448 
Puberty, albuminuria, of, 227 
Pulmonary actinomycosis, 45 
abscess, 80 
cancer, 83 
congestion, 83 
echinococcus disease, 1 1 , 5 1 
gangrene, 77 
hemorrhages, 58, 81 
infarction, 83 
lues, 84 
mycoses, 85 
saccharomycoses, 42 
tuberculosis, blood in, 635 
sputum in, 54 
Purin bases, 120 
Purulent gastritis, 370 

sputum, 3 
Pus casts, 267 

cells in prostatic fluid 

308 
formation, anaemia due to, 

599 
in gastric juice, 362 
in sputum, 11 
in stools, 395 
in urine, 274 
masses in sputum, 5 
Putrescin, 257 
Putrid bronchitis, sputum of, 

72 
Pycnosis, 503 
Pyelitis, acute, 291 
leucocytes in, 516 
productiva, 221 
urine in, 336 
Pyelonephritis, 336 

leucocytes in, 516 
Pyelonephrosis, leucocytes in, 

516 
Pyenotic nuclei, 503 
Pyloric stenosis, 365 
Pyogenic albumosuria, 236 
inflammations, leucocytes 
in, 514, 516 
Pyonephrosis, urine in, 338 
Pyrocatechin, 148 
Pyronin stain, 471 



Quadriurates, 116, 245 
Quartan malaria, parasite of, 

663 
Quotient, albumin, 217 

R 

Rabies, blood in, 514 
Rabinowitch's bacillus, 29 
Reaction of blood, 530 



Reaction of sputum, 3 
of stools, 387 
of urine, 100 
Rectum, cancer of, blood in. 
652 
stools in, 423 
Recurrent vomiting, 339 
Red cells, 445 

brassy cells, 447 
budding, 447 
color of, 447 
counting methods, 

452 
crenated, 446 
deformed, 446 
degenerations of, 448 
elasticity of, 447 
granules in, 447, 449 
in sputum, 14 
in urine, 237 
nucleated, 448 
nuclei fate of, 502 
number of, 445, 482 
origin of, 503 
poikilocytes, 446, 605 
resistance of, 489 
shape of, 445, 473 
size of, 447 
staining properties, 

475 
structure of, 473 
diarrhea, 423 
indigo, in urine, 147 
kidney, 331 
Reducible body of Stokvis, 150 
Reflex vomiting, 339 
Reichmann's disease, 368 
Relapsing fever, 677 
Renal atrophy, 335 

urine in, 335 
calculus, 338 

leucocytes in, 516 
concretions, 276 
diabetes, 165, 551 
diagnosis, 309 
disease, albuminuria, 232 
diuresis, 91 

blood in, 655 
epistaxis, 237 
epithelial cells in urine, 

261 
haematuria, 237 
haemophilia, 237 
permeability, 318 
phthisis, 336 
threshold, 550 
Rennin, 160, 356 
Residuale albumin urie, 230 
Residue of dried blood, 431 
Resinous acids, 211, 213 
Resistance of red blood, 489 
Retention of chlorides, 129 
Reticulated cells, 479 
Rhabditis stercoralis, 413 

strongyloides, 413 
Rheumatism, acute articular, 

blood in, 513, 644 
Rhizopoda, 401 
Rice bodies, 57 
Rice-water vomitus, 341 
Rickets, blood in, 658 
Rieder's cell, 505 
Riegel's meals, 342 
Ring body of Cabot, 477 
Rosaniline test, 318 
Rosenau's blood agar, 285 
Rosenbach's reaction, 155 

test, bile, 152 
Rot, grinder's, sputum of, 5 
Round worms, 53, 407 
Rowntree and Geraghty's test, 

319 
Rubner's test, glucose, 183 

lactose, 183 
Russell and Brodie's method, 

coagulation blood, 434 
Rusty sputum, 61 



Sabourand's medium, 50 
Saccharometer, 179 
Saccharomyces busse, 42 

hominis, 42 
Saccharomycosis, 42 
Sago granules in sputum, 13 

in stools, 393 
Sahli's desmoid reaction, 354 

hemometer, 441 
Salicylic acid test for gastric 
motility, 366 
of renal sufficiency, 
318 
Salkowski's method of deter- 
mining the alkalinity of 
blood, 537 
method for leucin, 256 
method for purin bases, 

121 
test for pentose, 185 
Salt and water test, 311 
Sand, gall, 398 

intestinal, 400 

Sarcinae in gastric contents 

360, 363 

in sputum, 42 

in stools, 419 

in urine, 305 

Sarcoma, blood in, 517, 652 

pleural fluid in, 704 
Saturation deficit, 348 
Scarlet fever, blood in, 633 
Scherer's, albumin determina- 
tion, 215 

test for inosite, 186 
for leucin, 256 
Schistocytes, 474 
Schistosoma haematobium in 
stools, 418 
in urine, 306 
japonicum, 413 
mansoni, 307 
Schizogony 659 
Schizont, 659 

Schlosing method for am- 
monia determination, 123 
Schmidt's method bilirubin in 

stools, 390 
Schmidt and Strassburger's 

diet, 386 
Schonbein Almen test, 241 
Schottilius enriching method, 

423 
Schutze's coarsely granular 
cells, 451 
finely granular cells, 451 
granular masses, 452, 525 
Scindi sore, 673 
Sclerosis (arterio-) of kidney, 

urine in, 332 
Scolices, 52 
Scurvy, blood in, 658 
Scybala, 388 
Secondary anaemia, 592 
Sedimentation of red blood- 
corpuscles, 431 
Sediments, urine, 243 
bilirubin, 253 
biurate, 245 
carbonates, 257 
cholesterol, 253 
cystin, 278 
gastric, 360 
haematoidin, 253 
haemoglobin, 253 
heteroalbumose, 253 
hippuric acid, 253 
indigo, 253 
leucin, 254 
melanin, 253 
mucous, 261 
organized, 244, 261 
oxalates, 278 
phosphates, 248 
scheme, 258 
tyrosin, 254 



INDEX 



723 



Sediments, unorganized, 244 
urates, 245 
uric acid, 245 
xanthin, 253 
Seidel's test for inosite, 186 
Sellard's test, reaction blood, 

204, 538 
Senile atrophy of kidney, 

urine in, 331, 335 
Septicaemia, blood in, 631 

urine in, 290 
"Seromucus sputum," 69 
Serous cysts, ovarian, 707 

pleurisy perforating 

through lung, 80 
Serous sputum, 3 

of oedema of lung, 80 
Serum albumin in urine, 218 
globulin in urine, 218 
specific gravity of, 430 
Sex, effect on count of red 

cells, 483 
Shape of red cells, 445, 473 
Shiga's bacillus, 424 
Siderosis, 5 

Silicic acid in urine, 140 
Skatoxyl, 147 
Skin, eosinophilia in diseases 

of, 523 
Sleeping sickness, 671, 675 
Small mononuclears, 494 

neutrophiles, 498 
Small pox, blood in, 634 
Smears, blood, 462 
Smegma bacillus, 29, 30 
bacilli in urine, 283 
Soaps in stools, 392 

determination, 392 
Sodium biurate, 246 
Sodium in urine, 142 
Soft chancre, organism oi, 300 
Specific gravity of blood, 429 
of body fluids, 693 
of urine, 92 
of plasma, 430 
of serum, 430 
of spinal fluid, 679 
Spermatocele, 709 
Spermatorrhoea, 273, 298 

anemia due to, 599 
Spermatozoa, 273, 308 
Spermin crystals in prostatic 

fluid, 309 
Spiegler's albumin reagent, 

214 
Spinal fluid, 678 

bacteriology of, 685 

cholin in, 683 

cytology of, 684 

globulin in, 681 

glucose in, 679 

in cerebral haemorrhage, 

688 
in chorea, 688 
in compression of cord, 

689 
in encephalitis, 689 
in diabetes, 688 
in epilepsy, 688 
in hydrocephalus, 688 
in locomotor ataxia, 685 
in lues, 691 

in mental disease, 688 
in meningitis, 689 
in paresis, 685 
in tumors of brain, 688 
in uremia, 688 
in meningismus, 689 
proteids in, 681 
saliva constituents, 683 
urea in, 680 

Wassermann test colloidal 
gold test, 685 

in poliomyelitis, 692 
Soirals, Curschmann's, 7 
Spirillum carteri, 677 

cholerae asiaticae, 422 
duttoni, 677 



Spirillum carteri, non patho- 
genic, 423 

Novvi, 67 7 

obermeieri, 677 

of Deneke, 422 

of Finkler and Prior, 423 

of Massea, 422 

of Metchnikoff, 423 

of Vincent, 41 
Schuylkiliensis, 423 
Spirochaeta buccalis, 300 

of mouth, 300 

of Obermeier, 677 
Spirochaete pallidum, 300 

refringens, 302 
Spleen, eosinophilia in dis- 
eases of, 522 
Splenic albuminuria, 230 

anaemia, 602 
Splenomegaly, red cells in, 

488 
Splenomylogenous leukemia, 

617 
Spodogenic splenic tumor, 504 
Spontaneous glycosuria, 162 
Spore stains, 282 
Sporoblast, 665 
Sporogone, 659 
Sporozoit, 665 
Sputum, 1 

abscess of lung, 78 

air in, 5 

amount, 2 

animal parasites of, 50 

asthma, 66 

aspirated foreign body, 

78 
bronchitis acute, 68 
chronic, 70 

bacteria in, 18 

bile-stained, 4 

blood in, 14 

bloody, 4 

bronchiectasis, 74 

bronchopneumonia, 65 

cavity of lung, 57 

cells of, 1 1 

character of, 3 

chemical analysis of, 53 

children's, 2 

chromogenic bacteria in, 
5, 19 

color of, 3 

congestion pastive of lung, 
83 

consistency of, 3 

crystals, 18 

cultures of, 18 

Curschmann's spirals, 7 

definition, 1 

diptheria, 85 

diseases, 54 

dust in, 4 

edema of lung, 81 

elastic tissue in, 14 

empyema, 80 

epithelial cells in, 11 

extraneous matter in, 11 

fibrinous structures in, 9 

foreign bodies in, 11 

gangrene, 77 

glanders, 66 

green, 4 

haemoglobin stained, 4 

infarction of lung, 83 

influenza, 65 

in Pneumonia, 60 

malignant disease of lung, 
83 

perforating serous pleu- 
risy, 80 

syphilis of lung, 84 

tuberculosis, 54 

Vincent's angina, 87 

Herzfehlerzellen in, 4 

jaundiced, 4 

layers, 5 

lung stones, 9 



Sputum, macroscopic, 6 

microscopic, 11 

morning, 2 

moulds in, 45 

mucoid, 3 

mucopurulent, 3 

myelin in, 2 

normal persons, 2 

odor of, 5 

plant parasites, 18 

pneumoliths, 10 

pneumonokonioses, 4 

prune- juice, 63 

purulent, 3 

reaction of, 3 

rusty, 61 

sarcinae in, 42 

serous, 3 

tissue fragments in, 6 

washing the, 19 

yeasts in, 42 
Sputum coctum, 69 

crudum, 68 

fundum peteus, 5 

globosum, 60 
Staining blood, 466 

casts, 272 

vital, 477 
Stains for bacteria, 280 

for blood, 466 

for capsules, 32 

for flagella, 282 

for worms, 426 
Staphylococcus epidermidis al« 
bus, 19 

pyogenes albus, 19 

pyogenes aureus, 19 
Starch, digestion of, 357 

in stools, 396 
Starvation, anemia due to, 597 
Steatorrhea, 425 
Stenosis of pylorus, 365 
Sterling's aniline gentian vio- 
let, 35 
Stippled cells, 480 
Stokvis reducible body, 151 

test, bile, 154 
Stolinkow's method of quanta 

tative albumin, 217 
Stomach acidity of, 344 

bacteria in, 362 1 

cancer of, 375 

blood in, 620 

contents, see Gastric Con- 
tents, 341 

dilated, 367 

fasting, 341 

gastric juice, 342 

hemorrhagic erosions, 374 

lactic acid in, 377 

lues of, 374 

motility of, 341, 364 

sarcinae in, 360, 363 

ulcer of, 373 
Stomatitis, 37 

ulceromembranous, 87 
Stomatomycosis, 42 
Stone in bladder, 276 

in kidney, 276 

in lung, 9 

in ureter, 338 
Stone-cutter's phthisis, 5 
Stones, gall, 398 

intestinal, 398 

lung, 9 

pancreatic, 398 

pseudo-gall, 398 
Stools, 382; alcoholic, 389 

albumin in, 396 

amoebae in, 401 

bacteria in, 387, 419 

bile pigments in,"389 

blood in, 393 

carbohydrates in, 396 
Stools, cestodes in, 415 

cholera, 422 

cholesterol in, 398 



724 



INDEX 



Stools, Charcot-Leyden crys- 
tals in, 398 

clay-colored, 398 

color of, 389 

concretions in, 398 

consistency of, 388 

constituents of, 387 

crystals in, 397 

curds in, 396 

diastase, 397 

dysentery, 423 

enteroliths in, 398 

epithelium cells in, 397 

examination of, 386 

fat in, 390 

fats estimation, 390 

fatty, 390 

ferments in, 397 

forms of, 387 

frequency of, 387 

gall sand in, 398 

gallstones, 398 

in amoebic dysentery, 423 

in Asiatic cholera, 422 

in disease, 420 

in pancreatic disease, 424 

in rectal diarrhoea, 423 

jn typhoid fever, 420 

intestinal parasites, 401 

intestinal sand, 398 

macroscopy, 398 

method of preserving, 426 

microscopy, 397 

milk curds in, 396 

mucus in, 392 

muscle-fibres in, 396 
normal 387 

pancreatic stones, 398 

parasites in, 401 

plant parasites, 418 

pseudo gall-stones in, 398 

protozoa in, 397, 401 

pus in, 395 

reaction of, 398 

rhizopoda in, 401 

round worms, 407 

soaps and fats, 392 

starch in, 396 

trematodes, 413 

trypsin in, 397 

tubercle bacilli in, 420 

tumor fragments in, 401 

yeasts in, 418 
Strauss test, lactic acid, 358 
Streptococcus cystitis, 293 

alpha homolysaus, 21 

buccalis, 21 

cystitis, 293 

equinus, 21 

erysipelatos, 20 

fecalis, 21 

group, 19, 569 

longus, 20 

mitior, 21 

mucosus capsulatus, 21 

pyogenes, 20 

saprophyticus, 21 

viridans, 21 
Streptothrix eppingeri, 31 

infections, 32 

pseudotuberculosa, 31 
Strongyloidal, 409 
Strongyloides intestinalis, 413 
anaemia due to, 600 
in sputum, 53 

stercoralis, 413 
Structural albuminuria, 225 
Subacidity in cancer of stom- 
ach, 377 
Subacute indurative pneumo- 
nia, sputum in, 65 

nephritis, urine in, 329 
Succinic acid, 696 
SulphEemoglobinaemia, 529 
Sulphates of urine, 136, 252 
easily split, 137 
ethereal, 136, 139 
of urine, neutral, 137 



Sulphates, total, 136, 139 

unoxodized, 136 
Sulphide of hydrogen in urine, 

140 
Sulphocyanic acid, 140 
Sulph-haemoglobinemia, 529 
Sulphur, granules, 45 

total, in urine, 138 
Summer diarrhoeas of children, 

647 
Sunlight, lack of, anaemia due 

to, 597 
Superacidity, 367 
Supersecretion, 367 
Suppurative nephritis, 335 

urine in, 336 
Surra, 671 

Sympathetic nervous system, 
eosinophilia in diseases of, 
524 
Syncytiem cells, 452 
Synovial fluids, 704 
Syphilis, albuminuria of, 329 
blood in, 652 
lung, 84 

nephritis in, 233 
of trachea and bronchi, 74 
of stomach, 374 
organism of, 300 
spinal fluid, 691 



Tabes dorsalis, spinal fluid in, 
685 

mesenterica, 391 
Taenia cucumerina, 417 
echinococcus, 51 
nana, 417 
saginata, 416 
solium, 416 
Tallqvist scale, 443 
Tanret's solution, 214 
Tape worms, 415 
Taurocholic acid, 155, 222 
Teichmann's test, 240 
Temperature, effect of, on 

count of reds, 484 
Terminal hematuria, 296 
Tertian malaria, 660 
Test meal, breakfast, 342 
Dock's, 342 
Ewald-Boas, 342 
Fischer, 342 
for intestinal exami- 
nation, 385 
renal, 312 
Riegel, 342 
Testicle, cancer of, blood in, 

651 
Testicular casts in urine, 273 
Tetanus, bacillus of, 293 
Tetragenus, micrococcus, in 

sputum, 40 
Therapeutic measures, effect 

of, on count of reds, 486 
Thermolytic albuminuria, 233 
Thermophilic organisms, 419 
Thionin stain, 469 
Thiosulphuric acid, 140 
Thoracentesis, cause of albu- 
minous expectoration, 81 
Thornapple crystals, 246 
Threads, mucous, in urine, 274 
Threshold, renal, 552 
Throat cultures, 85 
Thrush, 44 

Thymus and leukaemia, 627 
Tide, alkaline, 101 
Timothy hay bacillus, 29 
Tissue, elastic, in sputum, 18 
fragments in sputum, 6 

in urine, 274 
lung, in lung abscess, 78 
in gangrene, 77 
Titratable alkalinity of blood, 
530 



Toisson's fluid, 454 
Tolerance in diabetes, 202 

carbohydrate, 551 
Tonsillitis, leucocytes in, 514. 

634 
Topfer's method, 347 
Tophus, 710 

Total acidity, gastric, 345, 348 
of urine, 104 
organic acids in gastric 
juice, 359 
Toxaemic jaundice, 149 
Toxic vomiting, 339 
Trachea, lues of, 74 
Transitional leucocytes, 450. 

496 
Transudates, 700 
Traumatic albuminuria, 231 
Trematode worms in parasites. 

413 
Treponema calligyrum, 300, 
304 
macrodentium, 300, 303 
microdentium, 300, 303 
mucosum, 300, 303 
pallidum, 300 
Triacid stain, 471 
Trichina (Trichinella) spiralis, 
407, 678 
in blood, 678 
Trichiniasis, eosinophilia in, 

523 
Trichiuria, 412 
Trichiuris trichiura, 412 
Trichocephalus dispar, 412 
Trichomonas hominis, 362, 403, 
404 
intestinalis, 405 
pulmonalis, 51 
vaginalis, 51, 404 
in urine, 306 
Triple phosphate, 248 
in sputum, 18 
stain, 47 
Tripperfaden in urine, 273, 296 
Troje's marrow cell, 505 
Trommer's test, glucose, 166 
Tropaelin OO, 345 
Tropics, anaemia of, 598 

splenomegaly of, 673 
Trousseau's test, bile 154 
Trypanosomiadae, 406 
Trypanosoma brucei, 671 

gambiene, 671 , 
Trypanosomiasis, 671 
Trypsin, 382 

in stools, 397 
in urine, 160 
Tryptophan test, 380 
Tsetse fly disease, 671 
Tsuchiya's reagent, 216 
Tubercle bacilli, in cerebro- 
spinal fluid, 692 
in sputum, 22 
in stools, 420 
in urine, 283 
bacillus, 22 

animal inoculation, 
cultivation, 27 
staining, 25 
Tuberculin reaction, eosino- 
philia in, 523 
Tuberculosis, acute miliary, 
blood in, 637; sputum in, 55 
albuminuria in, 328 
adrenals, blood in, 639, 

659 
avian, 29 
blood in, 634 
bones and joints, blood in, 

638 
chronic of lungs, blood 

in, 635 
complement fixation for, 

586 
fish, 29 
inestine, 638 



INDEX 



725 



Tuberculosis, kidneys, blood in, 
638; urine in, 336 
lymph glands, blood in, 

629 
meningitis, blood in, 638 
of bladder, 291 
peritonitis, blood in, 638 
pulmonary, acute miliary, 
55 
bronchopneumonia, 

55 
chronic ulcerative, 56 
fibroid form, 55 
gastric contents in, 

351 
haemoptysical, 58 
sputum, 54 
Tuberculous cystitis, 292 
meningitis, 690 
pneumonia, 513 
Tubo-ovarian cysts, 708 
Tumor fragments in gastric 
juice, 341 
in stools, 401 
in urine, 274 
of brain, spinal fluid 
in, 688 
Tunnel workers' anaemia, 410 
Typhoid ' agglutination tests, 
564 
bacillus, 287 

in stools, 421 
fever, blood in, 628 
bronchitis of, 70 
pneumonia of, spu- 
tum of, 65 
stools in, 420 
Typhus diagnosticum, 498 

fever, blood in, 632 
Tyrosin, 696 

Tyrosin in body fluids, 695 
in sputum, 18 
in urine, 254, 255 

u 

Udransky's test, bile acid, 156 
Uffelmann's test, lactic acid, 

358 
Ulcer of duodenum, 373 

of stomach, 3 73 
Ulceration, chronic tubercu- 
lous, sputum in, 56 
Ulcerative endocarditis, 514 _ 
Ulceromembranous stomatitis, 

87 
Uncinaria americana, 409, 410 

duodenalis, 410 
Uncinariasis, anasmia in, 600 
Unilateral nephritis, 335 
Unorganized sediment, 244 
Unoxidized sulphur in urine, 

136 
Unripe cells of Grawitz, 500 
Uremia, 334, leucocytes in, 517 

urine in, 333 
Uramidoglycuronic acid, 206 
Urate calcium, 246 

casts, 268 

concretions, 276 

sediment, 245 
Urates, 116 
Urea, 109 

crystals, 709 

delayed excretion of, 311 

estimation, 112 

frost, 710 

in blood, 541 

in spinal fluid, 680 

isolation, 114 

properties. 111 

tests, 111 

urease methods, 112 
Ureamia and ocidosis, 204 

spinal fluid in, 688 
Ureama and ocidosis, 204 

spinal fluid in, 688 



Ureteral calculas, 338 
Ureteritis, membranous, 221 
Urethra, infection of, 294 
Urethritis, 295 

nonspecific, 297 
Uric acid, 114, 246 

concretions, 276 
crystals, 246 
determination of, 117 
diathesia, 102 
endogenous, 115 
exogenous, 115 
reagent, 545 
standard solution, 
119, 546 
Urine, 88 

acetone, 187 
acidity of, 103, 134 
acids of, 129 
albumose in, 235 
animal gum, 187 
alkapton bodies in, 207 
amount of, 89 
amylase in, 160 _ 
animal parasites in, 306 
bacteria, 279 
bases of, 129 

Bence Jones body in, 233 
bile in, 97 
blood in, 97, 237 _ 
cancer fragments in, 284 
carbohydrates in, 162 
casts in, 264 
collection of, 88 
color of, 94 
concretions in, 276 
cultures of, 284 
day and night, 90 
diacetic in, 192 
echinococcus hooklets in, 

306 
fat in, 209 
ferments of, 159 
filaria, embryos in, 306 
flagellates in, 306 
fragments of tissue in, 274 

tumor in, 274 
general appearance of, 100 
glucose in, 162 
glycogen, 186 
glycuronic acid, 206 
hemoglobin in, 238 
inorganic acids and bases, 

129 
inosite, 186 
isomaltose, 187 
laiose, 187 
levulose, 181 
maltose, 187 
mineral acidity, 103 
moulds in, 306 
nematode worms in, 307 
nitrogenous bodies in, 104 
ammonia, 122 
creatin, 128 
creatinin, 125 
oxyproteinic acid, 128 
purin bases, 120 
urea, 109 
uric acid, 114 
odor of, 99 
organic acidity, 104 
oxybutyric acid, 194 
pentose, 183 
pigments in, 143 
plant saprophytes in, 306 
preservation of, 88, 243 
in abscess of kidney, 336 
in acute nephritis, 327 
in atrophy of kidney, 335 
in chronic nephritis, 332 
in diabetes insipidus, 205 

mellitus, 198 
in congenital cystic kid- 
ney, 335 
in cancer of kidney, 336 
in hydro vephrosis, 338 
in pyelitis, 373 



Urine, in renal calculus, 338 

in subacute nephritis, 329 

in suppurative nephritis 
336 

in tuberculosis of kidney, 
336 

in unilateral nephritis, 335 

in uremia, 333 
Urine, pro eids in, 210, 217 

albumin, 218 

euglobulin, 219 

fibrinogen, 224 

globulin, 218 

Morner's body, 219 

mucin, 219 

nucleo albumin, 219 

nucleohiston, 223 

reaction of, 100 

sarcinse in, 305 

sediments, 243 

specific gravity of, 92 

total acidity, 103 

yeasts in, 305 
Urobilin, 95 

in blood, 529 

Urobilinemia, 96 
Urobilinogen, 95 
Urochrome, 95 
Uroerythrin, 95 
Uroleucinic, acid, 208 
Urometer, 92 
Uronephrosis, 338 
Urorhodin, 147 
Urorosein, 147 
Urorubin, 147 



Vacuolated red cells, 448 
Valeric acid in gastric con* 

tents, 359 
Vascular hyposthenuria, 312 
Vaughan's granules, 449 
Vaquez disease, 488 
Vegetable parasites in urine, 

306 
Vermiculus, 666 
Vesical calculi, 286 
Vicarious oxaluria, 250 
Vincent's angina, 87 

spirillum, 41 
Vinegar eel in urine, 307 
Viscosity of blood, 437 
Vital staining, 477 
Volume index, 491, 605 
Vomitus, 339 

bile in, 340 

blood in, 340 

fsecal, 341 

green, 340 

mucus in, 340 

pancreatic fluid in, 340 

reaction, 339 

rice-water, 341 

tumor fragments in, 341 

w 

Water and salt test, 311 
Wassermann reaction, 575 
Waxy casts, 266 
Weidel's test, purin bases, 121 
Welch's capsule stains, 33 
Weyl's test, creatinin, 126 
Whetstone crystals of uric 
acid, 246 
of xanthin, 253 
Whip worm, 412 
White cells, 450, 491 
diarrhoea, 423 
kidney, large, urine in, 

397 
of marrow, 504 
Whooping-cough, leucocytes 
in, 513 
organism of, 37 



726 



INDEX 



Whooping-cough, sputum of, 
37 

Widal test, 564 

Williamson's blood test, 648 

Wilson's stain, 468 

Winter cough sputum of, 7 1 

Women, leucocytes in, 511 
pregnant in, 510 

Worry, anemia due to, 597 

Worms, mounting, 426 

nematode, of urine, 307 
round, in stools, 407 

Wright's opsonic index, 567 

Wynn's roller. 453 



Xanthin bases in urine, 120, 
121 

crystals, 253 

stones, 278 
Xanthochromia, 679 
Xylose, 184 



Yeasts, ansemia due to, 601 
in blood, 602 
in gastric contents, 360, 363 



Yeasts, in sputum, 42 

in stools, 418 

in urine, 305 
Yellow atrophy, blood in, 657 
Yellow fever, anaemia in, 599 



Zenker's fluid, 33 
Ziehl-Neelsen, carbol-fuchsin, 

25 
Zygote, 665 



